State-dependent pudendal nerve stimulation for bladder control

ABSTRACT

The present disclosure provides a device for state-dependent pudendal nerve stimulation for bladder control in a subject and methods of making and using the same.

Bladder function is comprised of two phases: a filling phase (urinestorage) and a voiding phase (urine evacuation) and efficient bladderfunction involves control of these phases mediated by continence andmicturition reflexes accomplished through coordinated sympathetic,parasympathetic and somatic neural activity [Beckel and HolstegeNeurophysiology of the Lower Urinary Tract, in Urinary Tract (2011)Springer Berlin Heldelberg, 149-169]]. In bladder dysfunction (such asover-active bladder (OAB), underactive bladder (UAB) or urinaryretention), one or more of these functions is disrupted, leading tosymptoms including urinary urgency, frequency, urgency incontinence, andnocturia. These symptoms often fail to improve following pharmacologicaltreatment (Brindley et al. Br J Urol 46: 453-62, 1974).

Alternatives to pharmacological treatment of bladder dysfunctionincluding sacral nerve neuromodulation and posterior tibial nervestimulation have had only limited success (Mangera et al. & Siddiqui etal.). These approaches can require major surgical intervention orfrequent visits with a medical professional, thereby limiting thesuitability of such treatments in the majority of patients. Thus, thereis a need for therapies for bladder dysfunction.

Pudendal nerve stimulation is a promising therapeutic option fortreatment of OAB and bladder dysfunction symptoms, though it remainsunclear how to optimally stimulate the pudendal nerve to reduce thesymptoms of bladder dysfunction.

WO2017/066572 describes stimulation of the pudendal nerve with a highintensity electrical signal to improve bladder capacity.

SUMMARY OF THE INVENTION

The inventors have investigated electrical stimulation of the pudendalnerve, and the branches thereof, and have devised an apparatus, andmethods of using such an apparatus, which addresses the shortcomings ofprevious treatments for bladder dysfunction. In particular,WO2017/066572 describes high intensity stimulation of the pudendal nerveto increase bladder capacity. The inventors describe that low intensitystimulation of the pudendal nerve improves bladder capacity. This hasthe advantage that the battery life of any apparatus applying the signalis prolonged, thereby making the apparatus more efficient and providinggreater convenience for both the patient and clinicians.

Furthermore, the inventors have identified that the improvement inbladder capacity arising from low intensity stimulation of the pudendalnerve is prolonged—that is, it is carried over from one micturitioncycle to the next even in the absence of further stimulation. Thesurprising finding that low intensity pudendal nerve stimulationimproves bladder capacity for at least 1 or 2 micturition cycles afterstimulation has stopped means that stimulation does not need to beapplied every micturition cycle for treatment to be effective. Thisfurther extends the potential battery life of any apparatus applying thestimulation and greatly improves patient comfort.

Bladder function is comprised of two phases: a filling phase (urinestorage) and a voiding phase (urine evacuation). It is thereforedesirable to improve the function of both phases in a patientexperiencing bladder dysfunction. As described in the Examples, as wellas improving bladder capacity, pudendal nerve stimulation can improvevoiding efficiency. The approaches to improving voiding efficiencydescribed herein are applicable in scenarios in which a low intensitypudendal nerve stimulation has been used to improve bladder capacity,and also when a high intensity pudendal nerve stimulation has been usedto improve bladder capacity

Examples below demonstrate that electrical stimulation of the pudendalnerve during the bladder filling phase followed by termination ofstimulation at onset of voiding phase results in improved voidingefficiency compared to controls in which no stimulation has beenapplied. The Examples also demonstrated that voiding efficiency can befurther improved by application of a second electrical signal duringvoiding phase. In particular, application of a second electrical signalof higher frequency than the first electrical signal results in improvedvoiding efficiency. Similarly, application of a second electrical signalto motor fibres of the pudendal nerve in a burst pattern also results inimproved voiding efficiency.

Therefore, in a first aspect, the present disclosure provides anapparatus for stimulating neural activity in a pudendal nerve of asubject, the apparatus comprising, consisting of, or consistingessentially of : at least one primary electrode configured to apply afirst electrical signal to said nerve; and a controller coupled to saidprimary electrode(s) and controlling the first electrical signal to beapplied thereby, wherein said controller is configured to cause said atleast one primary electrode to apply said first electrical signal thatstimulates neural activity in the pudendal nerve to produce an increasein bladder capacity, wherein the first electrical signal comprises an ACwaveform having a frequency in the range of from 0.1-50 Hz, preferably1-50 Hz, and wherein the first electrical signal has an amplitude in therange from 0.05 T to 10 T.

In a further aspect, the present disclosure provides a method oftreating bladder dysfunction in a subject comprising, consisting of, orconsisting essentially of applying a first electrical signal to apudendal nerve of the subject, wherein the first electrical signalcomprises an AC waveform having a frequency in the range of from 1-50 Hzand wherein the first electrical signal has an amplitude in the range offrom 0.05 T to 10 T.

In some embodiments, the first signal comprises a “low intensity”signal. In some embodiments, the first electrical signal comprises anamplitude in the range from 0.05 T to <2.0 T. In some embodiments, thefirst electrical signal comprises an amplitude in the range from 0.3 Tto <2.0 T. In some embodiments, the first electrical signal comprises anamplitude in the range of from 1 T to 1.5 T. In other embodiments, thefirst electrical signal comprises an amplitude of 1 T. In yet otherembodiments, the first electrical signal comprises an amplitude of 1.5T.

In another embodiment, the first electrical signal comprises a frequencyin the range of from 1-20 Hz. In some embodiments, the first electricalsignal comprises a frequency of 10 Hz. In other embodiments, the firstelectrical signal comprises a frequency of 20 Hz.

In another embodiment, the first electrical signal is applied no morefrequently than alternate micturition cycles. In other embodiments, thefirst electrical signal is applied no more frequently than every thirdmicturition cycle.

In another embodiment, the first signal comprises a “high intensity”signal. In such embodiments, the first electrical signal comprises anamplitude in the range from 2 T to 10 T. In other embodiments, the firstelectrical signal comprises an amplitude in the range from 2 T to 4 T.In yet another embodiment, the first electrical signal comprises anamplitude comprising 3 T.

In some embodiments, the first electrical signal comprises an ACwaveform having a frequency in the range of from 0.1 to 50 Hz. In someembodiments, the first electrical signal comprises an AC waveform havinga frequency in the range of from 1 to 50 Hz. In some embodiments, thefirst electrical signal applied comprises an AC waveform having afrequency of 1 to 45 Hz. In yet other embodiments, the first electricalsignal applied comprises an AC waveform having a frequency in the rangeof from 2 to 40 Hz. In other embodiments, the first electrical signalcomprises a frequency in the range of 1 to 10 Hz. In one embodiment, thefirst electrical signal comprises a frequency of 10 Hz.

In yet another embodiment comprising either “high intensity” or “lowintensity” first electrical signals, the signal is applied to sensoryfibres of the pudendal nerve. In some embodiments, the signal is appliedto a sensory branch of the pudendal nerve, for example the dorsal nerveof the penis/clitoris (DNP, also known as the dorsal genital nerve). Insome embodiments of the apparatus of the invention, the at least oneprimary electrode is configured to apply the first electrical signal tosensory fibres of said pudendal nerve, and the controller is configuredto cause said at least one primary electrode to apply the firstelectrical signal that stimulates neural activity in sensory fibres ofthe pudendal nerve to produce an increase in bladder capacity.

In some embodiments, application of the first electronic signal isstopped at onset of a bladder voiding phase. In relation to theapparatus of the invention, in certain embodiments the controller causesthe first electronic signal to be stopped when onset of a voiding phaseis detected. In certain embodiments, the application of the first signaland no second signal provides a positive voiding effect in a malesubject and a negative voiding effect in a female subject.

As noted above, bladder function can be further improved by increasingvoiding efficiency through application of a second electrical signal.Therefore, in some embodiments of both “high intensity” and “lowintensity” first electrical signal embodiments, a second electricalsignal is applied to stimulate the pudendal nerve or branches thereof.In such embodiments, the second electrical signal can be applied by saidat least one primary electrode(s). In certain alternative embodiments,the apparatus comprises at least one secondary electrode coupled to thecontroller and the second electrical signal is applied by the at leastone secondary electrode(s), the controller controlling the signal to beapplied thereby.

In certain embodiments, the second electronic signal comprises an ACwaveform having a higher frequency than the first electronic signal.

In certain embodiments where a second electrical signal is to beapplied, the second electrical signal comprises an AC waveform having afrequency in the range of from 20-50 Hz. In certain embodiments, thesecond electrical signal comprises an AC waveform having a frequency of30-40 Hz, optionally 33 Hz or 40 Hz. In certain embodiments wherein thesecond electrical signal has a frequency higher than the firstelectrical signal, the second electrical signal has a frequency of 33Hz.

In some embodiments, the controller is configured to cause a secondelectrical signal to be applied, wherein said second electrical signalstimulates neural activity in the pudendal nerve to produce an increasein voiding efficiency and wherein the second electrical signal comprisesan AC waveform having a frequency in the range of from 20 to 50 Hz. Insome embodiments, the second electrical signal applied comprises an ACwaveform having a frequency of 20 to 45 Hz. In yet other embodiments,the second electrical signal comprises an AC waveform having a frequencyin the range of from 20 to about 40 Hz. In some embodiments, the secondelectrical signal comprises an AC waveform having a frequency of about33 Hz. In certain embodiments, the application of 33 Hz provides apositive voiding effect in a male subject and a negative voiding effectin a female subject.

In certain embodiments, the second electrical signal comprises an ACwaveform and is applied in a burst pattern. In such embodiments of theapparatus of the invention, the apparatus comprises at least onesecondary electrode coupled to the controller and the second electricalsignal is applied by the at least one secondary electrode(s), whereinthe controller is configured to cause the second electrical signal to beapplied in a burst pattern.

In certain embodiments, the burst pattern comprises a signal bursthaving a duration of from 50 ms to 1000 ms. In certain embodiments, theburst pattern comprises a signal burst having a duration of 100 ms.

In such embodiments, the burst pattern comprises of a signal bursthaving a duration from 50 ms to 1000 ms repeated at an interval of from0.125 s to 2 s. In certain preferred embodiments, the burst patterncomprises of a signal burst having a duration of 100 ms repeated at aninterval of 0.5 s.

In some embodiments the burst pattern consists of a signal burst havinga duration from 50 ms to 1000 ms repeated at an interval of from 0.125 sto 2 s. In some embodiments, the burst pattern consists of a signalburst having a duration from 50 ms to 1000 ms repeated at an interval offrom 0.2 s to 2 s. In certain preferred embodiments, the burst patternconsists of a signal burst having a duration of 100 ms repeated at aninterval of 0.5 seconds.

In certain embodiments wherein the second electrical signal is to beapplied in a burst pattern, the second electrical signal is to beapplied to motor fibres of the pudendal nerve, for example to a motorbranch of the pudendal nerve. In such embodiments of the apparatus ofthe invention, the at least one secondary electrode(s) is (are)configured to apply the second electrical signal to motor fibres of saidpudendal nerve, and the controller is configured to cause said at leastone secondary electrode to apply the second electrical signal thatstimulates neural activity in motor fibres of the pudendal nerve toproduce an increase in voiding efficiency.

In yet other embodiments, the second electrical signal applied in aburst pattern comprises an AC waveform having a frequency in the rangeof from 20 to 50 Hz. In some embodiments, the second electrical signalapplied in a burst pattern comprises an AC waveform having a frequencyof 20 to 45 Hz. In one embodiment, the second electrical signal appliedin a burst pattern comprises an AC waveform having a frequency of 40 Hz.

The second electrical signal applied in a burst pattern may be at afrequency 40 Hz applied in pulses at different intervals, for example0.125 seconds, 0.21 seconds and 0.5 seconds.

In yet other embodiments, the second electrical signal applied in aburst pattern comprises an AC waveform repeated at a frequency in therange of from 2 to 20 Hz. In some embodiments, the second electricalsignal applied in a burst pattern comprises an AC waveform repeated at afrequency in the range of 2 to 10 Hz. In one embodiment, the secondelectrical signal comprises an AC waveform repeated at a frequency of 10Hz. In another embodiment, the second electrical signal is repeated at afrequency of 2 Hz. In another embodiment, the second electric signal isrepeated at a frequency of 4.76 Hz. In another embodiment, the secondelectric signal is repeated at a frequency of 8 Hz.

In some embodiments where a second electrical signal is (to be) applied,the second electrical signal comprises an AC waveform having anamplitude in the range of 0.5-4 T. In some embodiments, the secondelectrical signal comprises an AC waveform having an amplitude in therange 1-3 T. In some embodiments, the second electrical signal comprisesan amplitude of about 3 T. In other embodiments wherein the secondelectrical signal is applied in a burst pattern, the second electricalsignal comprises an amplitude of 1 T. In certain embodiments wherein thesecond electrical signal has a frequency higher than the firstelectrical signal, the second electrical signal has an amplitude of 3 T.In certain embodiments wherein the second electrical signal is appliedin a burst pattern, the second electrical signal has an amplitude of1.8-2.3 T.

In a further aspect the invention provides a method of treating bladderdysfunction in a subject comprising: i. implanting in the subject anapparatus according to the first aspect; ii. positioning at least oneprimary electrode of the apparatus in signalling contact with a pudendalnerve of the subject and, when the apparatus comprises at least onesecondary electrode, positioning said at least one secondary electrodeof the apparatus in signalling contact with a pudendal nerve of thesubject; iii. activating the apparatus to apply an electrical signal tothe pudendal nerve of the subject as caused by the controller.

Apparatuses and methods according to the invention have the furtheradvantage that they can be used in conjunction with pharmaceuticaltherapies for bladder dysfunction to reduce symptoms.

Therefore, in a further aspect the invention provides a pharmaceuticalcomposition comprising a compound for treating bladder dysfunction, foruse in a method of treating bladder dysfunction in a subject, whereinthe method is a method according to the invention, the method furthercomprising the step of administering an effective amount of thepharmaceutical composition to the subject.

In a further aspect, the invention provides a pharmaceutical compositioncomprising a compound for treating bladder dysfunction, for use intreating bladder dysfunction in a subject, the subject having anapparatus according to the invention implanted.

In certain embodiments, the compound for treating bladder dysfunction isan antimuscarinic compound or a β-adrenergic receptor agonist,optionally a β3-adrenergic receptor agonist. In certain embodiments, theantimuscarinic compound is selected from darifenacin, hyoscyamine,oxybutynin, tolterodine, solifenacin, trospium, or fesoterodine. Incertain embodiments, the β3-adrenergic receptor agonist is mirabegron.

In a further aspect, the invention provides a neuromodulation systemcomprising a plurality of apparatuses according to the invention. Incertain embodiments, each apparatus is arranged to communicate with atleast one other apparatus in the system, optionally all apparatuses inthe system. In certain embodiments, the system further comprises aprocessor arranged to communicate with the apparatuses of the system.

In a preferred embodiment of all aspects of the invention, the subjectto be treated or for which the apparatus is to be used is a humansubject.

In certain embodiments the subject is a male subject. In certainembodiments the subject is a female subject.

In all aspects, unless specified otherwise, “pudendal nerve” refers tothe pudendal nerve and its branches. In certain embodiments, the firstand/or second electronic signal is (to be) applied to the motor branchof the pudendal nerve. In certain embodiments, the first and/or secondelectronic signal is (to be) applied to the sensory branch of thepudendal nerve, In certain embodiments, the first and/or secondelectronic signal is (to be) applied to dorsal nerve of thepenis/clitoris (DNP). In certain embodiments, the first and/or secondelectronic signal is (to be) applied to perineal nerve.

Another aspect of the present disclosure provides all that is disclosedand illustrated herein.

BRIEF DESCRIPTION OF FIGURES

FIG. 1—Schematic drawings showing how apparatuses, devices and methodsaccording to the invention can be put into effect.

FIG. 2—Experimental Setup

(A) Placement of the bladder catheter, nerve cuff, and electrodes forrecording external urethral sphincter (EUS) electromyogram (EMG). Twodifferent types of EMG electrodes were used (in different experiments),percutaneous wires (B) and flat metal contacts embedded in a siliconesubstrate (C) that was placed underneath the pubic bone. (D) Viewthrough the surgical microscope of the isolated sensory pudendal nervein the ischiorectal fossa. (E) View showing placement of the nerve cuffon the sensory pudendal nerve. (F) Side view of a nerve cuff showing theorientation of the incoming wires, as well as the tabs that are used toopen the cuff. (G) View of the inside of the nerve cuff after pullingthe cuff open at the tabs. The exposed portion of the contacts are nearthe outside of the cuff.

FIG. 3—Sensory pudendal nerve stimulation increased bladder capacity

(A)(B) Example traces from two experiments showing increases in bladdercapacity with increasing stimulation amplitude at 10 Hz. Traces arebladder pressure and EUS EMG. Dotted lines above the pressure tracesindicate periods of stimulation. (C) Reflex EMG responses to stimulus atamplitudes of 600 and 650 μA. A reflex response at 25 ms is present at650 μA, but not at 600 μA. Other amplitudes are reported as fractions ofthe stimulus reflex threshold (or 1 T). (D) Summary data showing changesin bladder capacity, relative to non-stimulation control trials, as afunction of stimulus amplitude and repetition frequency (mean±standarderror). Bladder capacity varied as a function of amplitude and frequency(p<0.001 for both amplitude and rate, two-way ANOVA) with nostatistically significant interaction term (n=9 for 0.3 T to 1 T, n=7for 1.5 T, 20 Hz, and n=6 for 1.5 T, 1 and 10 Hz). Increasing stimulusamplitude led to increased bladder capacities. Statistically significantincreases from 1 T to 1.5 T stimulation were observed for 1 and 20 Hz(p=0.01 and p=0.03, paired t-tests), but not for 10 Hz. The averagenormalized bladder capacity during 10 Hz stimulation was greater thanduring 20 Hz stimulation only at 0.6 T (p<0.001).

FIG. 4: Impact of sensory pudendal nerve stimulation on voidingefficiency

(A) Portion of a trial with stimulation at 1.5 T, 20 Hz illustratingstimulus termination following the first expulsion of urine. Theexpelled urine was collected and measured. If a second void occurredwithin twenty seconds of stimulus termination the urine was collectedseparately and measured (n=7 experiments). The volume from the firstmicturition event contributed to the normalized voiding efficiency.Values were normalized relative to non-stimulation control trials.Volumes from both the first micturition event and the second micturitionevent (if present) contributed to the normalized “reflex” voidingefficiency. (B) Summary data showing changes in voiding efficiency,relative to non-stimulation control trials, as a function of stimulusamplitude and repetition frequency (mean±standard error). Voidingefficiency varied as a function of amplitude and frequency (p<0.001 forboth amplitude and rate, two-way ANOVA) with no statisticallysignificant interaction term (n=7 for 0.3 T to 1 T, n=6 for 1.5 T, 20Hz, and n=5 for 1.5 T, 1 and 10 Hz). Increasing stimulation amplitudedecreased normalized voiding efficiency. This decrease was larger at 10and 20 Hz than 1 Hz (C) Waiting up to 20 seconds following stimulustermination elicited subsequent bladder contractions that resulted inthe expulsion of additional fluid. Despite the increase in expelledfluid, voiding efficiency remained reduced for 1 Hz (p=0.002) and 10 Hz(p<0.001) at 1.5 T (the amplitude which led to the greatest increase inbladder capacity). Statistical tests versus controls consisted oft-tests comparing distributions to a mean of 1.

FIG. 5: Stimulation of the sensory pudendal nerve leads to increases inbladder capacity (stimulation carryover effect)

(A) Bladder capacity values for each trial in an experiment. Circlesindicate the start of each trial. Black rectangles have been added tohighlight trials which show a stimulus carryover effect. (B) Sequentialtrials collected during a single experiment. Stimulation at 0.6 T, 20 Hz(trial 0) increased bladder capacity and initially disrupted coordinatedvoiding. On the subsequent trial (trial 1) the bladder capacity remainedincreased. On the subsequent and final trial of the series and theexperiment (trial 2) the bladder capacity decreased, but remainedelevated compared to the control trial (trial-1) conducted prior tostimulation. (C) Distributions of scaled bladder capacity on the threetrials following stimulation at 1 or 1.5 T and 10 or 20 Hz. Bladdercapacities were scaled such that 0% is the bladder capacity of the trialpreceding stimulation and 100% is the bladder capacity of thestimulation trial. Only groups of trials in which stimulation increasedbladder capacity by at least 20% relative to the preceding trial and inwhich at least three non-stimulation trials followed are included (n=15,48% of trials). Box plots were created using Matlab's boxplot( ) commandand the centre bars represent the median value, box edges are the 25thand 75th percentiles, and the whiskers extend to the most extreme datapoints which are not considered to be outliers.

FIG. 6: Sensory pudendal nerve stimulation increases bladder capacity inan intravesical PGE2 model of overactive bladder.

(A) Example traces showing a decrease in bladder capacity withintravesical PGE2 and increase in bladder capacity with sensory pudendalstimulation. (B) Bladder capacity decreased with PGE2 (p=0.02) andincreased (relative to PGE2) with stimulation (p=0.004) (n=9). (C)Voiding efficiency increased with PGE2 (p<0.001) and decreased (relativeto PGE2) with stimulation (p=0.05; p=0.01 with removal of the outlier).All stimulation was at 20 Hz with amplitudes ranging from 100 to 800 μA(mean 330±63 μA S.E.). Voided volume (to compute the voiding efficiency)consists of volume expelled during the first micturition event throughone minute following stimulus termination.

FIG. 7: Increases in bladder capacity and voiding efficiency fromstate-dependent stimulation—Experiment 1

(A) Example trial in which no stimulation occurred. Average bladdercapacity and voiding efficiency from this type of trial were 12.5 ml and28% respectively. (B) Stimulation at 3 T, 10 Hz (DNP) increased bladdercapacity and voiding efficiency (17.9 ml, 36%—averages). Thesestimulation parameters were also used for (C) and (D), but withdifferent stimulus parameters during the void (state-dependentstimulation). (C) Stimulus was terminated at void onset, furtherincreasing voiding efficiency (16.8 ml, 46%—averages). (D) Stimulationat 3 T, 33 Hz (DNP) during voiding which also had a higher voidingefficiency than stimulation throughout (18.2 ml, 46%).

FIG. 8: Increases in bladder capacity and voiding efficiency fromstate-dependent stimulation—Experiment 2

(A) Example trial without stimulation. Average bladder capacity andvoiding efficiency values from all such trials was 17.2 ml and 6%. (B)Stimulation throughout a trial at 3 T, 10 Hz (DNP) increased bladdercapacity but not voiding efficiency (43.3 ml, 4%,—averages). (C) Same asB, but with termination of the stimulus at void onset. These trials hadincreased voiding efficiency relative to stimulation throughout (43.9ml, 17%—averages). (D) Same as B, but with 3 T, 33 Hz stimulation (DNP)during voiding. These trials also had increased voiding efficiency (47.6ml, 19%—averages). (E) Same as B, but with burst stimulation of pudendalmotor nerve during voiding, which led to increased voiding efficiency(50.8 ml, 83%—averages).

FIG. 9: Increases in bladder capacity and voiding efficiency fromstate-dependent stimulation—Experiment 3

(A) Example trial without stimulation. Average bladder capacity andvoiding efficiency values were 37.2 ml and 41%. (B) Stimulationthroughout (3 T, 10 Hz, DNP) increased bladder capacity slightly (40.8ml) with a decrease in voiding efficiency (20%). (C) Termination of thestimulus at void onset did not improve voiding efficiency (39.9 ml,22%—averages), nor did switching to stimulation at 33 Hz (D) (38.3 ml,14%—averages). (E) Motor burst stimulation increased voiding efficiency(40.8 ml, 78%—averages).

FIG. 10: Experimental setup and lower urinary tract anatomy.

A) Neuroanatomy of the rat lower urinary tract, including the sacralplexus and pudendal nerve (modified from McKenna and Nadelhaft, 1986).The inset shows this neuroanatomy relative to the pelvic bones andspinal column. The highlighted circle (in inset) shows our surgicalaccess point in the ischiorectal fossa. Grey boxes indicate placement ofnerve cuffs on the sensory branch as well as on the motor branch. B)Photo of the isolated sensory branch. A CorTec cuff has been placed nextto, but not on the nerve (modified from Hokanson et al., 2017a). C)Neuroanatomy of the male cat lower urinary tract (modified from Martinet al., 1974). Dorsal genital nerve cuff was placed proximal tosplitting of ischiourethralis branch. In females a nerve cuff was placedproximal to the splitting of the cranial sensory branch. D) A nerve cuff(CorTec) in the open position with stimulation sites exposed. The onlypoint of exposure of the two stimulation sites for this model are at theedges of the cuff, the rest of the metal is insulated. E) Side view ofthe same nerve cuff in the closed position showing the 300 μm diameteropening. F) Two of the three “bursting” stimulation patterns employedfor this study. Each tick represents a biphasic stimulation pulse. Inall trials (rat and cat) the burst consisted of 3 pulses at 40 Hz. Thetime between bursts varied depending on trial type and consisted of0.125 (8 Hz, rat), 0.21 (4.76, rat), or 0.5 s (2 Hz, rat and cat)between burst starts.

FIG. 11: Continuous stimulation of the sensory pudendal nerve in thefemale rat increased bladder capacity but decreased voiding efficiency.

A) Example cystometrograms from a single rat. Increasing stimulationamplitude increased bladder capacity as evident by the increase in filltime. The magnitude of bladder contraction decreased with increasingamplitude, which resulted in decreased voiding efficiency. Stimulationcontinued through the first contraction/leak event. A second contractionwas also recorded without stimulation, but these second contractionswere insufficient to increase voiding efficiency back to baseline. B)Summary bladder capacity data at 1, 10, and 20 Hz. Increasingstimulation amplitude increased bladder capacity, with the largestincreases at 10 Hz. C) Summary voiding efficiency data at 1, 10, and 20Hz. Increasing stimulation amplitude decreased voiding efficiency. Forsummary data n=9 for 0.3 T to 1 T, n=7 for 1.5 T, 20 Hz, and n=6 for 1.5T, 1 and 10 H.

FIG. 12: State-dependent stimulation in male cats.

A) Example cystometrograms without stimulation, with continuousstimulation, and with state-dependent stimulation. An inhibitorystimulus (10 Hz, 3 T) was used to promote bladder filling (inhibitbladder contractions). Two different excitatory stimuli, 33 Hzstimulation and motor bursting, were used to promote bladder emptying.Continuous stimulation increased bladder capacity but decreased voidingefficiency by 24%. B) State-dependent stimulation increased bladdercapacity (n=6) relative to control trials. C) State-dependentstimulation increased voiding efficiency relative to continuousstimulation (p=0.003 for ANOVA test, p=0.028 for fill only, p=0.048 for33 Hz, p<0.001 for motor bursting, n=5); in all experiments motorbursting increased voiding efficiency relative to control trials.Bladder capacities from all stimulation trials were averaged acrossexperiments as the inhibitory stimulus was always 3 T, 10 Hz on thedorsal genital nerve (DGN).

FIG. 13: State-dependent stimulation in the female rat (n=9) usingelectrical stimulation of the sensory pudendal nerve (10 Hz, 1 T) toinhibit bladder contractions (promote bladder filling) and eitherstimulus termination (fill-only condition) or motor bursting patterns topromote bladder emptying. A) Example cystometrograms from a singleexperiment. Lines below the EUS EMG traces demonstrate the duration ofsensory pudendal stimulation. Lines above the EUS EMG traces demonstratethe duration of motor bursting stimulation. Values on the left areaverage bladder capacity and voiding efficiency data for trials of thistype in this experiment. B) Normalized bladder capacity for allstimulation trials relative to controls. All relevant trials within anexperiment were averaged together resulting in a single data point foreach experiment. C) Normalized voiding efficiency for all stimulationconditions relative to controls. State-dependent stimulation increasedvoiding efficiency relative to continuous stimulation (p=0.023 forANOVA, p=0.016 for fill-only, p=0.001 for 2 Hz, p=0.003 for 4.76 Hz,p=0.004 for 8 Hz, n=5).

FIG. 14: State-dependent stimulation in a female cat and side-by-sidesummary data from male and female cats. A) Example cystometrogramswithout stimulation, with continuous stimulation, and withstate-dependent stimulation. Continuous stimulation increased bladdercapacity but decreased voiding efficiency by 52% (Voiding efficiency wascalculated as voided volume divided by the voided and residual volumes).B) State-dependent stimulation increased bladder capacity (n=6 males, 5females) relative to control trials. C) State-dependent stimulationincreased voiding efficiency relative to continuous stimulation; in allexperiments motor bursting increased voiding efficiency relative tocontrol trials. Experiment counts were n=6M,5F (M=male, F=female) forcontinuous stimulation, n=6M,4F for fill only stimulation, n=6M,4F for33 Hz stimulation, n=5M,5F for motor bursting stimulation. Bladdercapacities from all stimulation trials were averaged across experimentsas the inhibitory stimulus was always 3 T, 10 Hz on the dorsal genitalnerve (males) or sensory pudendal nerve (females).

DETAILED DESCRIPTION OF THE INVENTION

For the purposes of promoting an understanding of the principles of thepresent disclosure, reference will now be made to preferred embodimentsand specific language will be used to describe the same. It willnevertheless be understood that no limitation of the scope of thedisclosure is thereby intended, such alteration and furthermodifications of the disclosure as illustrated herein, beingcontemplated as would normally occur to one skilled in the art to whichthe disclosure relates.

The terms as used herein are given their conventional definition in theart as understood by the skilled person, unless otherwise defined below.In the case of any inconsistency or doubt, the definition as providedherein should take precedence.

Articles “a” and “an” are used herein to refer to one or to more thanone (i.e. at least one) of the grammatical object of the article. By wayof example, “an element” means at least one element and can include morethan one element.

“About” is used to provide flexibility to a numerical range endpoint byproviding that a given value may be “slightly above” or “slightly below”the endpoint without affecting the desired result.

The use herein of the terms “including,” “comprising,” or “having,” andvariations thereof, is meant to encompass the elements listed thereafterand equivalents thereof as well as additional elements. Embodimentsrecited as “including,” “comprising” or “having” certain elements arealso contemplated as “consisting essentially of” and “consisting of”those certain elements.

Recitation of ranges of values herein are merely intended to serve as ashorthand method of referring individually to each separate valuefalling within the range, unless otherwise-Indicated herein, and eachseparate value is incorporated into the specification as if it wereindividually recited herein. For example, if a frequency range is statedas 1 Hz to 50 Hz, it is intended that values such as 2 Hz to 40 Hz, 10Hz to 30 Hz, or 1 Hz to 3 Hz, etc., are expressly enumerated in thisspecification. These are only examples of what is specifically intended,and all possible combinations of numerical values between and includingthe lowest value and the highest value enumerated are to be consideredto be expressly stated in this disclosure.

As used herein, the term “subject” and “patient” are usedinterchangeably herein and refer to both human and nonhuman animals. Theterm “nonhuman animals” of the disclosure includes all vertebrates,e.g., mammals and non-mammals, such as nonhuman primates, sheep, dog,cat, rat, horse, cow, chickens, amphibians, reptiles, and the like.

As used herein, “electrode” is taken to mean any element capable ofapplying an electrical signal to the nerve.

As used herein, “stimulation of neural activity” may be an increase inthe total signalling activity of the whole nerve, or that the totalsignalling activity of a subset of nerve fibres of the nerve isincreased, compared to baseline neural activity in that part of thenerve. A “selective increase in neural activity”, for example in thesensory fibres, causes a preferential increase in neural activity in thesensory fibres compared to any increase in neural signalling in themotor nerve fibres of the pudendal nerve. By way of further example, aselective increase in neural activity of the motor fibres causes apreferential increase in neural activity in the motor fibres compared toany increase in neural signalling in the sensory nerve fibres of thepudendal nerve.

“Phase-specific” or “state-dependent” stimulation are taken to mean thata different stimulation is applied depending on the ongoing and/ordesired phase of the normal bladder activity cycle. The bladder activitycycle or micturition cycle is characterised by a filling phase (alsoreferred to as a storage phase), followed by a triggering of themicturition, followed by a voiding phase (also referred to as themicturition phase). A normal bladder activity cycle is a bladderactivity cycle characteristic of a healthy individual.

Application of an electrical signal in a “burst pattern” refers toapplication of the signal in a series of bursts. That is, the signal isapplied for a burst—that is, a duration of time—followed by an intervalin which no signal is applied. The interval is then followed by thesignal being again applied for another burst, followed by anotherinterval. The burst pattern is the combination of the burst for whichthe signal is applied followed by the interval during which no intervalis applied.

The “ongoing phase” of bladder activity is the phase of the bladderactivity cycle occurring at a particular given time. That a subject isin a given phase of the cycle can be indicated by a physiologicalparameter relevant to bladder activity, for example bladder pressure.For example, that a subject is in the filling phase may be indicated byincreasing bladder pressure, or a sustained bladder pressure indicatingthat the bladder is at least partially filled. Triggering of micturitionmay be indicated by a sharp increase in bladder pressure. Otherphysiological parameters relevant to bladder activity include nerveactivity in the pudendal nerve, nerve activity in the hypogastric nerve,nerve activity in the pelvic nerve, muscle activity in the bladderdetrusor muscle, muscle activity in the internal urethral sphincter,muscle activity in the external urethral sphincter (EUS), muscleactivity in the external anal sphincter (EAS).

The “desired phase” of the bladder activity cycle is the phase of thebladder activity cycle of which the subject is desirous. The desiredphase may depend on the behaviour of the subject, for example whetherthey are sleeping, at exercise, at work, etc. Similarly, the desiredphase may depend on perceived levels of urinary comfort. For example,the subject may perceive discomfort due to the sensation of having afull bladder, and therefore be desirous of triggering micturition.

It will be appreciated that phase-specific stimulation can take intoaccount both ongoing and desirous phases of the bladder activity cycle.For example, a first stimulating signal may be applied (e.g. to increasebladder capacity) during a filling phase indicated by increasing bladderpressure, and a second stimulating signal may be applied when thesubject is desirous of beginning micturition (e.g. to triggermicturition), or during a voiding phase as indicated by a change inmuscle activity in the EUS (e.g. to increase voiding efficiency).

As used herein “pudendal nerve” refers to the compound pudendal nerveand its associated branches, for example the dorsal nerve of thepenis/clitoris (DNP) or the motor branch of the pudendal nerve.

As used herein, a “healthy individual” or “healthy subject” is anindividual not exhibiting any disruption or perturbation of normalbladder activity.

As used herein, “bladder dysfunction” is taken to mean that the patientor subject is exhibiting disruption of bladder function compared to ahealthy individual. Bladder dysfunction may be characterised by symptomssuch as nocturia, increased urinary retention, increased incontinence,increased urgency of urination or increased frequency of urinationcompared to a healthy individual. Bladder dysfunction includesconditions such as overactive bladder (OAB), neurogenic bladder, stressincontinence, underactive bladder (UAB), and urinary retention.

Treatment of bladder dysfunction, as used herein may be characterised byany one or more of a reduction in number of incontinence episodes, adecrease in urgency of urination, a decrease in frequency of urination,an increase bladder capacity, an increase in bladder voiding efficiency,a decrease in urinary retention, a change in external urethral sphincter(EUS) activity towards that of a healthy individual, and/or a change inthe pattern of action potentials or activity of the pudendal nervetowards that of a healthy individual.

The skilled person will appreciate that the baseline for any neuralactivity or physiological parameter in an individual need not be a fixedor specific value, but rather can fluctuate within a normal range or maybe an average value with associated error and confidence intervals.Suitable methods for determining baseline values would be well known tothe skilled person.

As used herein, a measurable physiological parameter is detected in asubject when the value for that parameter exhibited by the subject atthe time of detection is determined. A detector is any element able tomake such a determination.

A “predefined threshold value” for a physiological parameter is thevalue for that parameter where that value or beyond must be exhibited bya subject or subject before the intervention is applied. For any givenparameter, the threshold value may be defined as a value indicative of apathological state (e.g. the subject is experiencing abnormal retentionof urine) or a particular physiological state (e.g. the subject having afull bladder), or a particular behavioural state (e.g. the subjectwishes to begin voiding/micturition). Examples of such predefinedthreshold values include: bladder pressure abnormal compared to ahealthy individual, bladder pressure indicative of bladder at or nearcapacity, abnormal peripheral nerve activity (for example, pudendalnerve, hypogastric nerve or pelvic nerve) compared to a healthyindividual, abnormal EUS activity compared to a healthy individual (forinstance an increase in EUS activity). Such a threshold value for agiven physiological parameter is exceeded if the value exhibited by thesubject is beyond the threshold value—that is, the exhibited value is agreater departure from the normal or healthy value for that parameterthan the predefined threshold value.

The measurable physiological parameter may comprise an action potentialor pattern of action potentials in one or more nerves of the subject,wherein the action potential or pattern of action potentials isassociated with bladder dysfunction. Suitable nerves in which to detectan action potential or pattern of action potentials include a pudendalnerve, a pelvic nerve and/or a hypogastric nerve. In a particularembodiment, the measurable physiological parameter comprises the patternof action potentials in the pudendal nerve.

The measurable physiological parameter may be muscle electromyographicactivity, wherein the electromyographic activity is indicative of thelevel of activity in the muscle. Such activity could typically bemeasured from the bladder detrusor muscle, the internal urethralsphincter, the external urethral sphincter, and the external analsphincter.

As used herein, “implanted” is taken to mean positioned within thesubject's body. Partial implantation means that only part of theapparatus is implanted—i.e. only part of the apparatus is positionedwithin the subject's body, with other elements of the apparatus externalto the subject's body. Wholly implanted means that the entire apparatusis positioned within the subject's body. For the avoidance of doubt, theapparatus being “wholly implanted” does not preclude additionalelements, independent of the apparatus but in practice useful for itsfunctioning (for example, a remote wireless charging unit or a remotewireless manual override unit), being independently formed and externalto the subject's body.

In accordance with a first aspect of the invention, there is provided anapparatus for stimulating neural activity in a pudendal nerve of asubject, the apparatus comprising: at least one primary electrodeconfigured to apply a first electrical signal to said nerve; and acontroller coupled to said primary electrode(s) and controlling thefirst electrical signal to be applied thereby, wherein said controlleris configured to cause said at least one primary electrode to apply saidfirst electrical signal that stimulates neural activity in the pudendalnerve to produce an increase in bladder capacity, wherein the firstelectrical signal comprises an AC waveform having a frequency in therange of from 0.1-50 Hz and wherein the first electrical signal has anamplitude in the range 0.05 to 10 T. In certain embodiments the firstelectrical signal has an amplitude in the range 0.1 to 10 T.

“T” is a measure of relative stimulation intensity. Relative stimulationintensity can be expressed as multiples (0.1, 0.8, 1, 2, 5, etc.) of“T”. “T” represents the threshold stimulation intensity to evoke a motorresponse. For example, “1 T” is defined as the threshold stimulationintensity required to evoke a motor response—in particular, as usedherein “T” may be defined as the threshold amplitude required to evoke areflex electromyogram (EMG) response in the external urethral sphincter(EUS) when the electrical signal is applied to the pudendal nerve. Asanother example, the term “1 T” may be defined at the thresholdamplitude required to evoke a reflex EMG response in the external analsphincter (EAS). The type of motor response measured may dependent onthe subject. For example, in rats, the EUS is measured. In cats, the EASis measured. By way of further example, “1 T” may be defined in humansas the threshold amplitude required to evoke a perceptible sensation.

Determining “T” as described herein provides a calibration baseline ableto be transferred between individuals and/or species (e.g., rats vs.cats). T determined by either EUS or EAS provides a useful measure foramplitude normalization between individuals and/or species. For example,T may be determined as follows: a low frequency electrical signal (e.g.,1 Hz) is applied and the intensity of stimulation is increased (eitherby increasing the voltage or the current of the signal, preferably thecurrent) until the pudendal nerve stimulation produces a reflex EMGresponse in the EUS (or EAS). This stimulation intensity is designatedT. The absolute threshold stimulation intensity may vary acrossindividuals and/or species due to inherent variation, positioning andtype of the electrode, etc., and therefore subsequent experimental ortherapeutic intensities are designated as multiples of T to provideequivalent relative stimulation intensities.

The desired stimulation intensity (i.e. the desired multiple ofthreshold intensity “T”) can be achieved through controlled variation ofthe current or voltage of the signal, preferably the current.

In some embodiments the first electrical signal is a “low intensity”signal. In such embodiments, the first electrical signal has anamplitude in the range from 0.05 T to <2.0 T. In additional embodiment,the first signal has an amplitude in the range from 0.3 T to <2 T. Incertain embodiments, the first electrical signal has an amplitude valueof from 0.5 T to 1.5 T.

In some embodiments, the first electrical signal has an amplitude of0.05, 0.1, 0.2, 0.3, 0.4, 0.5 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3,1.4, 1.5, 1.6, 1.7, 1.8, 1.9, or 2.0 T. In some embodiments, theelectrical signal has a T value of 1 T-1.5 T. In other embodiments, thefirst electrical signal has an amplitude of 1 T. In other embodiments,the first electrical signal has an amplitude of 1.5 T.

In other embodiments the first electrical signal is a “high intensity”signal. In such embodiments, the first electrical signal has anamplitude in the range from 2 T to 10 T. In some embodiments, the firstelectrical signal comprises has an amplitude value of from 2 T to 4 T.In such embodiments, the first electrical signal has an amplitude of2.0, 2.1, 2.2, 2.3, 2.4, 2.5, 2.6, 2.7, 2.8, 2.9, 3.0, 3.1, 3.2, 3.3,3.4, 3.5, 3.6, 3.7, 3.8, 3.9 or 4.0 T. In certain embodiments, the firstelectrical signal has an amplitude of 3 T.

In other embodiments, the first electrical signal has an amplitude inthe range of from 0.1 to 20 mA. In certain embodiments, the firstelectrical signal has an amplitude in the range of from 0.1-10 mA,optionally 0.1-5 mA, optionally 0.1-1 mA, optionally 100-500 μA,optionally 100-400 μA. In certain embodiments, the first electricalsignal has an amplitude of 100 μA, 200μA, or 400 μA.

In certain embodiments, the apparatus may comprise two or more primaryelectrodes, where each primary electrode is configured to apply thefirst electronic signal. In certain such embodiments, the apparatuscomprises two primary electrodes suitable for bilateral positioning.

In certain embodiments, the first electrical signal stimulates neuralactivity in sensory fibres of the pudendal nerve, optionally selectivelystimulates neural activity in sensory fibres of the pudendal nerve, soas to produce an increase in bladder capacity.

In certain embodiments, the first electronic signal comprises an ACwaveform having a frequency in the range of from 1-50 Hz. In certainembodiments the first electronic signal comprises an AC waveform havinga frequency in the range of 1-20 Hz, for example 1, 2, 3, 4, 5, 6, 7, 8,9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 Hz. In certain embodimentswherein the first signal is a “low intensity” signal, the firstelectronic signal has a frequency of 10 or 20 Hz. In certain embodimentswherein the first signal is a “high intensity” signal, the firstelectronic signal has a frequency of 10 Hz.

It will be appreciated by the skilled person that an electronic signal“comprising” an indicated frequency may have other frequency componentsas part of the signal. In certain preferred embodiments of all aspectswhere a signal comprises an indicated frequency, the indicated frequencyis the dominant frequency component of the signal.

In another embodiment, the controller is configured to cause a secondelectrical signal to be applied, wherein said second electrical signalstimulates neural activity in the pudendal nerve to produce an increasein voiding efficiency. In some embodiments the second electrical signalcomprises an AC waveform having a frequency higher than the frequency ofthe first electrical signal.

In certain such embodiments, the second signal has a frequency in therange of from 5-50 Hz, optionally 20-40 Hz. In certain embodiments, thesecond signal has a frequency of 20-50 Hz. In certain embodiments, thesecond signal has a frequency of 30-40 Hz Hz. In certain embodiments,the second electrical signal has a frequency of 33 Hz. In certainembodiments, the second electrical signal has a frequency of 40 Hz. Incertain such embodiments, the second electronic signal stimulates neuralactivity in sensory fibres of the pudendal nerve.

In some embodiments, the second electronic signal is applied by the atleast one primary electrodes. In certain alternative embodiments, thesecond electrical signal is applied by at least one secondaryelectrode(s) coupled to said controller, said controller controlling thesignal to be applied thereby.

In another embodiment, the apparatus further comprises at least onesecondary electrode configured to apply a second electrical signal tosaid nerve and coupled to said controller, said controller controllingthe signal to be applied thereby, wherein said controller is configuredto cause said secondary electrode to apply said second electrical signalthat stimulates neural activity in the pudendal nerve to produce anincrease in voiding efficiency, wherein the second electrical signalcomprises an AC waveform and wherein said controller is configured tocause said second electrical signal to be applied in a burst pattern.

In certain embodiments, the burst pattern comprises a signal bursthaving a duration of from 50 ms to 1000 ms. In certain embodiments, theburst pattern comprises a signal burst having a duration of 100 ms.

In certain embodiments, the burst pattern comprises a signal bursthaving a duration from 50 ms to 1000 ms repeated at an interval of from0.125 s to 2 s. In certain embodiments, the burst pattern comprises asignal burst having a duration from 50 ms to 1000 ms repeated at aninterval of from 0.2 s to 2 s. In certain embodiments, the burst patterncomprises a signal burst having a duration of 100 ms repeated at aninterval of 0.5 s.

In certain embodiments, the burst pattern consists of a signal bursthaving a duration from 50 ms to 1000 ms repeated at an interval of from0.125 s to 2 s. In certain embodiments, the burst pattern consists of asignal burst having a duration from 50 ms to 1000 ms repeated at aninterval of from 0.2 s to 2 s. In certain embodiments, the burst patternconsists of a signal burst having a duration of 100 ms repeated at aninterval of 0.5 s.

In some embodiments, the second electrical signal applied in a burstpattern comprises a signal burst repeated at a frequency in the range offrom 0.5 to 20 Hz. For example, a burst patent comprising a signal burstrepeated at a frequency of 2 Hz would repeat the signal burst at 0.5 sintervals (FIG. 10F). In some embodiments, the second electrical signalapplied in a burst pattern comprises a signal burst repeated at afrequency in the range of 2 to 20 Hz. In some embodiments, the secondelectrical signal applied in a burst pattern comprises a signal burstrepeated at a frequency in the range of 2 to 10 Hz. In such embodimentsthe signal burst comprises an AC waveform.

In one embodiment, the second electrical signal comprises a signal burstrepeated at a frequency comprising 10 Hz, optionally wherein the secondelectrical signal comprises an AC waveform repeated at a frequency of 10Hz. In another embodiment, the second electrical signal is repeated at afrequency comprising 2 Hz, optionally wherein the second electricalsignal is repeated at a frequency of 2 Hz. In another embodiment, thesecond electrical signal is repeated at a frequency comprising 4.76 Hz,optionally wherein the second electric signal is repeated at a frequencyof 4.76 Hz. In another embodiment, the second electrical signal isrepeated at a frequency comprising 8 Hz, optionally wherein the secondelectric signal is repeated at a frequency of 8 Hz.

In certain embodiments, the second electrical signal applied in a burstpattern comprises a signal burst wherein the signal burst comprises anAC waveform having a frequency in the range of from 20-50 Hz. In certainembodiments, the signal burst comprises an AC waveform having afrequency of 30-40 Hz. In certain such embodiments, the signal burst hasa frequency of 40 Hz.

In some embodiments, the burst pattern consists of from 1 to 10 pulsesper signal burst. In some embodiments, the burst pattern consists offrom 1 to 5 pulses per signal burst. In some embodiments, the burstpattern consists of 3 pulses per signal burst. In such embodiments, theduration of the signal burst is thus determined by the frequency of theAC waveform.

In other embodiments where a second electrical signal is (to be)applied, the second electrical signal comprises an AC waveform having anamplitude in the range of 0.5-4 T. In some embodiments, the secondelectrical signal comprises an AC waveform having an amplitude in therange 1-3 T. In some embodiments, the second electrical signal comprisesan amplitude of about 3 T. In other embodiments wherein the secondelectrical signal is applied in a burst pattern, the second electricalsignal comprises an amplitude of 1 T. In other embodiments where thesecond electrical signal is applied in a burst pattern, the secondelectrical signal comprises an amplitude of 1.8-2.3 T.

In certain embodiments, the second electrical signal has an amplitude of0.1-20 mA, optionally 0.1-10 mA, optionally 0.1-5 mA, optionally 0.1-1mA, optionally 100-500 μA, optionally 100-400 μA. In certainembodiments, the second electrical signal has an amplitude of 100 μA,200μA, or 400 μA.

In other embodiments, the second electrical signal stimulates neuralactivity in motor fibres of the pudendal nerve, optionally selectivelystimulates neural activity in motor fibres of the pudendal nerve, so asto produce an increase in voiding efficiency. In such embodiments, thesecondary electrode(s) may be configured to be positioned on a motorbranch of the pudendal nerve.

In certain embodiments, the apparatus may comprise two or more secondaryelectrodes, where each primary electrode is configured to apply thesecond electronic signal. In certain such embodiments, the apparatuscomprises two secondary electrodes suitable for bilateral positioning.

The following embodiments apply equally to the first and secondelectronic signals unless specified otherwise.

In certain embodiments, the AC waveform is a biphasic waveform,optionally a charge-balanced biphasic waveform. In certain suchembodiments, the waveform may be symmetrical or asymmetrical. In certainsuch embodiments, each phase of the biphasic waveform has a phaseduration from 0.005 ms to 2 ms, optionally 0.01 to 1 ms, optionally 0.05to 0.5 ms, optionally 0.05 to 0.2 ms, optionally 0.1 ms. In certainembodiments, each phase of a biphasic waveform is of equal duration. Incertain alternative embodiments, each phase is of a different duration.

The AC waveform may be selected from sinusoidal, triangular, square or acomplex waveform.

In certain embodiments, the apparatus further comprises a detector todetect one or more physiological parameters in the subject. Such adetector may be configured to detect one physiological parameter or aplurality of physiological parameters The detected physiologicalparameter(s) are selected from nerve activity in the pudendal nerve,nerve activity in the hypogastric nerve, nerve activity in the pelvicnerve, muscle activity in the bladder detrusor muscle, muscle activityin the internal urethral sphincter, muscle activity in the externalurethral sphincter, muscle activity in the external anal sphincter, andbladder pressure.

In such embodiments, the controller is coupled to the detectorconfigured to detect a physiological parameter and causes the controllerto cause the first electrical signal to be applied when thephysiological parameter is detected to be meeting or exceeding a firstpredefined threshold value, for example when the detected valueindicates that the subject is in the filling phase of the micturitioncycle. In certain embodiments, the controller causes the firstelectronic signal to be stopped when the detector detects onset of abladder voiding phase.

Where a second electronic signal is to be applied, the detector maycause the controller to cause the second electrical signal to be appliedwhen a physiological parameter is detected to be meeting or exceeding asecond predefined threshold value, for example when the detected valueindicates that the subject is in the voiding phase of the micturitioncycle.

It will be appreciated that any two or more of the indicatedphysiological parameters may be detected in parallel or consecutively.For example, in certain embodiments, the controller is coupled to adetector or detectors configured to detect the pattern of actionpotentials in the pudendal nerve at the same time as the bladderpressure in the subject.

In addition, or as an alternative to a detector, the apparatus maycomprise an input element. In such embodiments, the input element allowsthe subject to enter data regarding their behaviour and/or desires. Forexample, the input element may allow the subject to enter that theydesire to begin bladder voiding (i.e. intend to begin urinating). Insuch embodiments, the controller is configured to cause a signal to beapplied that produces a physiological response appropriate to the datainput—for example, in the case of the intention to urinate beingindicated, the signal may increase voiding efficiency. By way of furtherexample, the input element may also allow the subject to enter dataindicative of behaviour in which storage phase is appropriate (e.g.sleeping or following urination, where it is desirous to promotestorage). In response to such data being entered via the input element,the controller causes a signal to be applied that produces aphysiological response appropriate for improved storage, for exampleincreased bladder capacity. The input element may be connected directlyto the controller, or be in wireless communication as a remotecomponent, for example a component carried by the subject. Sucharrangements and configurations are discussed in further detail below.

In certain embodiments, the apparatus further comprises one or morepower supply elements, for example a battery, and/or one or morecommunication elements.

In certain embodiments, the apparatus is suitable for at least partialimplantation into the subject, optionally full implantation into thesubject.

In a second aspect, the present disclosure provides a method of treatingbladder dysfunction in a subject comprising:

i. implanting in the subject an apparatus according to the invention;

ii. positioning at least one primary electrode of the apparatus insignalling contact with a pudendal nerve of the subject and, when theapparatus comprises at least one secondary electrode, positioning saidat least one secondary electrode of the apparatus in signalling contactwith a pudendal nerve of the subject;

iii. activating the apparatus to apply an electrical signal to thepudendal nerve of the subject as caused by the controller. The apparatusis activated when the apparatus is in an operating state such that thesignal will be applied as determined by the controller.

In certain embodiments the first signal is applied during a fillingphase and the second signal applied during a voiding phase. In certainembodiments, application of the first electronic signal results in anincrease in bladder capacity. In certain embodiments in which theapparatus is configured to apply a second electronic signal, applicationof the second electronic signal results in an increase in voidingefficiency.

In certain embodiments, the method comprises implanting an apparatusaccording to the invention having at least two primary electrodes, andoptionally at least two secondary electrodes, and positioning theelectrodes bilaterally—that is, one primary electrode in signallingcontact with the left pudendal nerve, and one primary electrode insignalling contact with the right pudendal nerve.

In certain embodiments, the method is a method for treating overactivebladder, neurogenic bladder, mixed urge and stress incontinence, underactive bladder (UAB), urinary retention, or detrusor hyperactivity withimpaired contractility (DHIC).

Implementation of all aspects of the present disclosure (as discussedboth above and below) will be further appreciated by reference to FIGS.1A-1C.

FIGS. 1A-1C show how the invention may be put into effect using one ormore apparatuses which are implanted in, located on, or otherwisedisposed with respect to a subject in order to carry out any of thevarious methods described herein. In this way, one or more apparatusescan be used to treat bladder dysfunction in a subject, by stimulatingneural activity in a pudendal nerve.

In FIG. 2A a separate apparatus 100 is provided for unilateralneuromodulation, although as discussed above and below an apparatuscould be provided for bilateral neuromodulation (100, FIG. 1B and 10).Each such apparatus may be fully or partially implanted in the subject,or otherwise located, so as to provide neuromodulation of the respectivenerve or nerves. FIG. 1A also schematically shows in the cutawaycomponents of one of the apparatuses 100, in which the apparatuscomprises several elements, components or functions grouped together ina single unit and implanted in the subject. A first such element is anelectrode 102 which is shown in proximity to a pudendal nerve 90 of thesubject. The apparatus may optionally further comprise furtherelectrodes (not shown) implanted proximally to the same or otherpudendal nerve. Alternatively, the other pudendal nerve may be providedwith a separate apparatus 100 (not shown). The primary electrode 102 maybe operated by a controller 104. The apparatus may comprise one or morefurther elements such as a communication element 106, a detector 108, apower supply element 110 and so forth. Each apparatus 100 may operateindependently, or may operate in communication with each other, forexample using respective communication elements 106.

Each neuromodulation apparatus 100 may carry out the requiredstimulation in response to one or more control signals. Such a controlsignal may be provided by the controller 104 according to an algorithmindependently, in response to output of one or more detector elements108, and/or in response to communications from one or more externalsources (for example an input element) received using the communicationselement. As discussed herein, the detector(s) could be responsive to avariety of different physiological parameters.

FIG. 1B illustrates some ways in which the apparatus of FIG. 1A may bedifferently distributed. For example, in FIG. 1B the apparatuses 100comprise electrodes 102 implanted proximally to a pudendal nerve 90, butother elements such as a controller 104, a communication element 106 anda power supply 110 are implemented in a separate control unit 130 whichmay also be implanted in, or carried by the subject. The control unit130 then controls the electrodes in both of the apparatuses viaconnections 132 which may for example comprise electrical wires and/oroptical fibres for delivering signals and/or power to the electrodes.

In the arrangement of FIG. 1B one or more detectors 108 are locatedseparately from the control unit, although one or more such detectorscould also or instead be located within the control unit 130 and/or inone or both of the apparatuses 100. The detectors may be used to detectone or more physiological parameters of the subject, and the controlleror control unit then causes the transducers to apply the first or secondsignal in response to the detected parameter(s), for example only when adetected physiological parameter meets or exceeds a predefined thresholdvalue. Physiological parameters which could be detected for suchpurposes may be selected from nerve activity in the pudendal nerve,nerve activity in the hypogastric nerve, nerve activity in the pelvicnerve, muscle activity in the bladder detrusor muscle, muscle activityin the internal urethral sphincter, muscle activity in the externalurethral sphincter, muscle activity in the external anal sphincter, andbladder pressure.

A variety of other ways in which the various functional elements couldbe located and grouped into the neuromodulation apparatuses, a controlunit 130 and elsewhere are of course possible. For example, one or moresensors of FIG. 1B could be used in the arrangement of FIG. 1A or 1C orother arrangements.

FIG. 1C illustrates some ways in which some functionality of theapparatus of FIG. 1A or 1B is provided not implanted in the subject. Forexample, in FIG. 10 an external power supply 140 is provided which canprovide power to implanted elements of the apparatus in ways familiar tothe skilled person, and an external controller 150 provides part or allof the functionality of the controller 104, and/or provides otheraspects of control of the apparatus, and/or provides data readout fromthe apparatus, and/or provides a data input element 152. The data inputfacility could be used by a subject or other operator in various ways,for example to input data relating to the behaviour of the subjectand/or their desires (e.g. that they are in a voiding phase).

By way of further example, devices for stimulating nerve activity in thepudendal nerve are described in U.S. Pat. Nos. 7,571,000 and 8,396,555,each of which are incorporated herein by reference.

In a further aspect, the invention provides a method of treating bladderdysfunction in a subject comprising applying a first electrical signalto a pudendal nerve of the subject, wherein the first electrical signalcomprises an AC waveform having a frequency in the range of from 0.1-50Hz and wherein the first electrical signal has an amplitude in the rangefrom 0.05 T to 10 T. In certain embodiments, the first electrical signalhas an amplitude in the range from 0.1 T to 10 T.

In other embodiments, application of the method is mediated using anapparatus according to the present disclosure.

In another embodiment, first electrical signal is applied to thepudendal nerve during a bladder filling phase, wherein application ofsaid first electrical signal increases bladder capacity.

In another embodiment, application of the first electrical signal isstopped at the onset of a bladder voiding phase. Where the method isperformed by an apparatus as described herein, such halting of the firstelectrical signal may be caused by voiding being detected by theapparatus or may be caused by the subject indicating via an inputelement that voiding phase had commenced.

In yet another embodiment, the first electrical signal stimulates neuralactivity in sensory fibres of the pudendal nerve, optionally selectivelystimulates neural activity in sensory fibres of the pudendal nerve, soas to produce an increase in bladder capacity.

In some embodiments the first electronic signal comprises an AC waveformhaving a frequency 1 Hz to about 50 Hz. In some embodiments, the firstelectrical signal applied comprises an AC waveform having a frequency of1 to about 45 Hz. In yet other embodiments, the first electrical signalapplied comprises an AC waveform having a frequency in the range of from20 to about 40 Hz. In other embodiments, the first electrical signalcomprises a frequency in the range of 2 to about 10 Hz.

In certain embodiments, the application of a second electrical signal,which has a frequency 33 Hz, provides a positive voiding effect (i.e.,increased voiding efficiency) in a male subject and a negative voidingeffect (i.e., decreased voiding efficiency) in a female subject. Incertain embodiments, the application of the first signal and no secondsignal provides a positive voiding effect in a male subject and anegative voiding effect in a female subject

In another embodiment, the method further comprises applying a secondelectrical signal to a pudendal nerve of the subject, wherein the secondelectrical signal comprises an AC waveform and is applied in a burstpattern.

In certain embodiments, the burst pattern comprises a signal bursthaving a duration of from 50 ms to 1000 ms. In certain embodiments, theburst pattern comprises a signal burst having a duration of 100 ms.

In some embodiments, the second electrical signal applied in a burstpattern comprises a signal burst repeated at a frequency in the range offrom 0.5 to 20 Hz. For example, a burst patent comprising a signal burstrepeated at a frequency of 2 Hz would repeat the signal burst at 0.5 sintervals (FIG. 10F). In some embodiments, the second electrical signalapplied in a burst pattern comprises a signal burst repeated at afrequency in the range of 2 to 20 Hz. In some embodiments, the secondelectrical signal applied in a burst pattern comprises a signal burstrepeated at a frequency in the range of 2 to 10 Hz. In such embodiments,the signal burst comprises an AC waveform.

In some embodiments, the burst pattern consists of a signal burst havinga duration from 50 ms to 1000 ms repeated at an interval of from 0.125 sto 2 s. In some embodiments, the burst pattern consists of a signalburst having a duration from 50 ms to 1000 ms repeated at an interval offrom 0.2 s to 2 s. In certain embodiments, the burst pattern consists ofa signal burst having a duration of 100 ms repeated at an interval of0.5 s.

In some embodiments, the second electrical signal applied in a burstpattern comprises an AC waveform repeated at a frequency in the range offrom 0.5 to 20 Hz. In some embodiments, the second electrical signalapplied in a burst pattern comprises an AC waveform repeated at afrequency in the range of 2 to 20 Hz. In some embodiments, the secondelectrical signal applied in a burst pattern comprises an AC waveformrepeated at a frequency in the range of 2 to 10 Hz.

In one embodiment, the second electrical signal is applied in a burstpattern comprising an AC waveform repeated at a frequency comprising 10Hz, optionally wherein the second electrical signal comprises an ACwaveform repeated at a frequency of 10 Hz. In another embodiment, thesecond electrical signal is repeated at a frequency comprising 2 Hz,optionally wherein the second electrical signal is repeated at afrequency of 2 Hz. In another embodiment, the second electrical signalis repeated at a frequency comprising 4.76 Hz, optionally wherein thesecond electric signal is repeated at a frequency of 4.76 Hz. In anotherembodiment, the second electrical signal is repeated at a frequencycomprising 8 Hz, optionally wherein the second electric signal isrepeated at a frequency of 8 Hz.

In some embodiments, the second electrical signal applied in a burstpattern comprises a signal burst comprising an AC waveform having afrequency in the range of from 20-50 Hz. In certain embodiments, thesecond electrical signal applied in a burst pattern comprises a signalburst comprising an AC waveform having a frequency of 30-40 Hz. Incertain such embodiments, the second electronic signal has a frequencyof 40 Hz.

The second electrical signal applied in a burst pattern may be at afrequency of 40 Hz applied in bursts at different intervals, for example0.125 seconds, 0.21 seconds and 0.5 seconds. In another embodiment, thesecond electrical signal is repeated at a frequency of 2 Hz. In anotherembodiment, the second electric signal is repeated at a frequency of4.76 Hz. In another embodiment, the second electric signal is repeatedat a frequency of 8 Hz.

In some embodiments, the burst pattern consists of from 1 to 10 pulsesper signal burst. In some embodiments, the burst pattern consists offrom 1 to 5 pulses per signal burst. In some embodiments, the burstpattern consists of 3 pulses per signal burst. In such embodiments, theduration of the signal burst is thus dictated by the frequency of the ACwaveform.

In another embodiment the second electrical signal has an amplitudevalue of from 0.5 T to 4 T, optionally 1-3 T. In certain embodimentswherein the second electrical signal has a frequency higher than thefirst electrical signal, the second electrical signal has an amplitudeof 3 T. In certain embodiments wherein the second electrical signal isapplied in a burst pattern, the second electrical signal has anamplitude of 1.8-2.3 T.

In some embodiments, the second electrical signal has an amplitude of0.1-20 mA, optionally 0.1-10 mA, optionally 0.1-5 mA, optionally 0.1-1mA, optionally 100-500 μA, optionally 100-400 μA. In certainembodiments, the second electrical signal has an amplitude of 100 μA,200μA, or 400 μA.

In another embodiment, the second electrical signal stimulates neuralactivity in motor fibres of the pudendal nerve, optionally selectivelystimulates neural activity in motor fibres of the pudendal nerve, so asto produce an increase in voiding efficiency. In certain suchembodiments, the second electrical signal is applied to a motor branchof the pudendal nerve.

The following embodiments apply equally to the first and secondelectronic signals unless specified otherwise.

In certain embodiments, the AC waveform is a biphasic waveform,optionally a charge-balanced biphasic waveform. In certain suchembodiments, the waveform may be symmetrical or asymmetrical. In certainsuch embodiments, each phase of the biphasic waveform has a phaseduration from 0.005 ms to 2 ms, optionally 0.01 to 1 ms, optionally 0.05to 0.5 ms, optionally 0.05 to 0.2 ms, optionally 0.1 ms. In certainembodiments, each phase of a biphasic waveform is of equal duration. Incertain alternative embodiments, each phase is of a different duration.

The AC waveform may be selected from sinusoidal, triangular, square or acomplex waveform.

In certain embodiments, treatment of bladder dysfunction, for exampleoveractive bladder, may be characterised by a combination of an increasein bladder capacity during filling periods and an increase in voidingefficiency for voiding periods.

In certain embodiments, the method is a method for treating overactivebladder, neurogenic bladder, underactive bladder (UAB), urinaryretention, or detrusor hyperactivity with impaired contractility (DHIC).

As demonstrated in the Examples, stimulation of the pudendal nerve inaccordance with the invention can improve bladder function duringsubsequent micturition cycles, even when stimulation is no longer beingapplied. This presents a significant advantage, as therapeutic benefitcan be achieved across multiple micturition cycles as a result ofstimulation in only one. This results in reduced power demand and lessimpact on the patient without a loss of therapeutic benefit. Therefore,in certain embodiments, the first electronic signal is applied no morefrequently than alternate micturition cycles—that is, no more frequentlythan every two micturition cycles—for example, every 2, every 3, orevery 4 micturition cycles. In certain embodiments, the first electronicsignal is applied no more frequently than every three micturitioncycles. In certain embodiments the first electronic signal is appliedevery two micturition cycles—i.e. every other filling phase. By way ofexplanation, in such embodiments, the first electronic signal is appliedduring the filling phase of one cycle. The signal may be then stoppedfor the voiding phase of that cycle and is not applied during the nextfilling phase or voiding phase—i.e. for a complete cycle. Once acomplete micturition cycle has occurred with no first electronic signalbeing applied, the first electronic signal is then applied during thesubsequent filling phase.

In a further aspect the invention provides a neuromodulation system, thesystem comprising a plurality of apparatuses according to the invention.In such a system, each apparatus may be arranged to communicate with atleast one other apparatus, optionally all apparatuses in the system. Incertain embodiments, the system is arranged such that, in use, theapparatuses are positioned to bilaterally stimulate the pudendal nervesof a patient.

In such embodiments, the system may further comprise additionalcomponents arranged to communicate with the apparatuses of the system,for example a processor, a data input facility, and/or a data displaymodule. In certain such embodiments, the system further comprises aprocessor. In certain such embodiments, the processor is comprisedwithin a mobile device (for example a smart phone) or computer.

In a further aspect, the invention provides a pharmaceutical compositioncomprising a compound for treating bladder dysfunction, for use in amethod of treating bladder dysfunction in a subject, wherein the methodis a method according to the invention, the method further comprisingthe step of administering an effective amount of the pharmaceuticalcomposition to the subject. It is a preferred embodiment that thepharmaceutical composition is for use in a method of treating bladderdysfunction wherein the method comprises applying a signal to a part orall of a pudendal nerve of said patient to stimulate the neural activityof said nerve in the patient, the signal being applied by aneuromodulation apparatus.

In a further aspect, the invention provides a pharmaceutical compositioncomprising a compound for treating bladder dysfunction, for use intreating bladder dysfunction in a subject, the subject having anapparatus according to the invention implanted. That is, thepharmaceutical composition is for use in treating a subject that has hadan apparatus as described according to the first aspect implanted. Theskilled person will appreciate that the apparatus has been implanted ina manner suitable for the apparatus to operate as described. Use of sucha pharmaceutical composition in a patient having an apparatus accordingto the first aspect implanted will be particularly effective as itpermits a cumulative or synergistic effect as a result of thecombination of the compound for treating bladder dysfunction andapparatus operating in combination.

In certain embodiments of these aspects, the compound for treatingbladder dysfunction is selected from an antimuscarinic compound and aβ-adrenergic receptor agonist, optionally a β3-adrenergic receptoragonist. In certain embodiments, the antimuscarinic compound is selectedfrom darifenacin, hyoscyamine, oxybutynin, tolterodine, solifenacin,trospium, or fesoterodine. In certain embodiments, the β-adrenergicreceptor agonist is a β3-adrenergic receptor agonist, for examplemirabegron. In certain embodiments, the pharmaceutical composition isfor use in treating OAB.

In certain embodiments, the pharmaceutical composition may comprise apharmaceutical carrier and, dispersed therein, a therapeuticallyeffective amount of the compounds for treating bladder dysfunction. Thecomposition may be solid or liquid. The pharmaceutical carrier isgenerally chosen based on the type of administration being used and thepharmaceutical carrier may for example be solid or liquid. The compoundsof the invention may be in the same phase or in a different phase thanthe pharmaceutical carrier.

Pharmaceutical compositions may be formulated according to theirparticular use and purpose by mixing, for example, excipient, bindingagent, lubricant, disintegrating agent, coating material, emulsifier,suspending agent, solvent, stabilizer, absorption enhancer and/orointment base. The composition may be suitable for oral, injectable,rectal or topical administration.

For example, the pharmaceutical composition may be administered orally,such as in the form of tablets, coated tablets, hard or soft gelatinecapsules, solutions, emulsions, or suspensions. Administration can alsobe carried out rectally, for example using suppositories, locally orpercutaneously, for example using ointments, creams, gels or solution,or parenterally, for example using injectable solutions.

For the preparation of tablets, coated tablets or hard gelatinecapsules, the compounds for treating bladder dysfunction may be admixedwith pharmaceutically inert, inorganic or organic excipients. Examplesof suitable excipients include lactose, maize starch or derivativesthereof, talc or stearic acid or salts thereof. Suitable excipients foruse with soft gelatine capsules include, for example, vegetable oils,waxes, fats and semi-solid or liquid polyols.

For the preparation of solutions and syrups, excipients include, forexample, water, polyols, saccharose, invert sugar and glucose. Forinjectable solutions, excipients include, for example, water, alcohols,polyols, glycerine and vegetable oil. For suppositories and for localand percutaneous application, excipients include, for example, naturalor hardened oils, waxes, fats and semi-solid or liquid polyols.

The pharmaceutical compositions may also contain preserving agents,solublizing agents, stabilizing agents, wetting agents, emulsifiers,sweeteners, colorants, odorants, buffers, coating agents and/orantioxidants.

Thus, a pharmaceutical formulation for oral administration may, forexample, be granule, tablet, sugar coated tablet, capsule, pill,suspension or emulsion. For parenteral injection for, for example,intravenous, intramuscular or subcutaneous use, a sterile aqueoussolution may be provided that may contain other substances including,for example, salts and/or glucose to make to solution isotonic. Thecompound may also be administered in the form of a suppository orpessary or may be applied topically in the form of a lotion, solution,cream, ointment or dusting powder.

In a preferred embodiment of all aspects of the invention, the subjector subject is a mammal, more preferably a human. In certain embodiments,the subject or subject is suffering from bladder dysfunction, forexample OAB.

The foregoing detailed description has been provided by way ofexplanation and illustration and is not intended to limit the scope ofthe appended claims. Many variations in the presently preferredembodiments illustrated herein will be apparent to one of ordinary skillin the art and remain within the scope of the appended claims and theirequivalents.

EXAMPLES

I. Experimental Dataset 1

The following experimental methods were used throughout Examples 1-5discussed below.

Surgical Preparation and Equipment Setup

Female Wistar rats (n=18) weighing between 237 and 296 g wereanesthetized with urethane (1.2 g/kg SC, supplemented as necessary).Body temperature was monitored using an esophageal temperature probe andmaintained at 36-38° C. with a water blanket. Heart rate and arterialblood oxygen saturation levels were monitored using a pulse oximeter(Nonin Medical Inc., 2500 A VET).

For cystometrogram (CMG) measurements, the bladder was exposed via amidline abdominal incision. A PE-90 catheter, the tip of which washeated to create a collar, was inserted into the bladder lumen through asmall incision in the apex of the bladder dome and secured with a 6-0silk suture. The abdominal wall was closed in layers with 3-0 silksuture. The bladder catheter was connected via a 3-way stopcock to aninfusion pump (Braintree Scientific Inc., BS-8000 or Harvard ApparatusPHD 4400) and to a pressure transducer (ArgoTrans, ArgonMedical DevicesInc., Plano, Tex.) connected to a bridge amplifier and filter(13-6615-50, Gould Instruments, Valley View, Ohio) for measuringintravesical pressure (IVP). Data were sampled at 1 kHz using a PowerLabsystem (AD Instruments, Colorado Springs, Colo.).

External urethral sphincter (EUS) EMG was measured using two differentapproaches (FIG. 2A). Experiments with Prostaglandin E2 (PGE2) used twoPFA-coated platinum-iridium wires (140 μm diameter, A-M Systems, Sequim,Wash.). These wires were inserted percutaneously using a needle topierce the skin, one on each side of the urinary meatus (FIG. 2B).Saline-only experiments used two platinum contacts bonded to a siliconebacking with wires welded to each contact (FIG. 2C). Thissheet-electrode was placed between the urethra and the pubic symphysisusing an intra-abdominal approach. EUS EMG leads were connected througha preamplifier (HIPS, Grass Products, Warwick, R.I.) to an amplifier(P511, Grass Products), and a subcutaneous needle served as ground.Signals were filtered (3 Hz-3 kHz) and sampled at 20 kHz.

After placing the bladder catheter and EUS EMG electrodes, the animalwas carefully flipped into a prone position. After resecting glutealmuscles at the midline, the ischium was spread apart from the sacrum toexpose the ischiorectal fossa, and the sensory branch of the pudendalnerve was isolated from connective tissue (FIG. 2D). For PGE2experiments custom bipolar nerve cuffs were placed around the sensorybranch and held in place with the use of Kwik-Cast (World PrecisionInstruments, Sarasota, Fla.). The custom cuffs consisted of 10 strandedstainless steel wire (AS631, Cooner Wire Co, Chatsworth, Calif.) placedthrough 2 mm long, 500 μm inner-diameter, silicone tubing approximately1 mm apart. The exposed wires were insulated with silicone (MED-1137,NuSil, Carpinteria, Calif.) and a slit was cut along the length of thetubing to allow placement of the nerve into the cuff. For saline-onlyexperiments, the same custom nerve cuff was used in one experiment. Forother saline-only experiments nerve cuffs that were 200 μm innerdiameter with 2 mm length (n=4) or 300 μm inner diameter with 3 mmlength (n=4) (FIG. 2F,G) (CorTec, Freiburg, Germany) were used.Following nerve cuff placement the opening was sutured closed in layerswith 3-0 silk suture. The animal was then carefully flipped back into asupine position for cystometric testing.

Preliminary testing indicated that 100 μM PGE2 consistently reducedbladder capacity as a model of OAB. PGE2 (Sigma-Aldrich) was dissolvedin ethanol to 10 mM concentration and stored in a −20° C. Freezer. Ondays of experiments, this stock solution was diluted in saline to thedesired concentration.

Experimental Procedures

The stimulus amplitude required to evoke a reflex response in theexternal urethral sphincter (EUS) was determined. With the bladderempty, stimuli were delivered at 0.1 to 1 Hz to the sensory pudendalnerve while monitoring evoked EUS EMG activity on an oscilloscope. Theamplitude required to evoke consistently (more than 50% of the time)reflex activation of the EUS was considered the stimulus threshold or 1T (1 times threshold) and was verified to within 10% accuracy.Experiments with percutaneous EMG electrodes (PGE2 experiments) did notshow consistent reflex activation of the sphincter. In these experimentsa 200 μA stimulus was used primarily (n=6), based on some preliminarytesting with intravesical saline.

The bladder was filled continuously with physiological saline at roomtemperature (2-8 ml/hr) using an infusion pump with an open urethra forat least 45 minutes during post-surgical recovery. The bladder wassubsequently emptied and cystometrograms (CMGs) recorded. For each CMG,the bladder was filled until a micturition event was observed, at whichtime the infusion pump was turned off. For PGE2 experiments,approximately one minute after the bladder pressure returned tobaseline, the bladder was emptied via the catheter using a syringe. Fornon-stimulated trials in saline-only experiments the bladder was emptiedimmediately following bladder pressure return to baseline. Forstimulation in saline-only experiments, two approaches were used. In thefirst two experiments, the bladder was emptied upon pressure return tobaseline, the same as during non-stimulated trials. In the last sevenexperiments, the stimulus was terminated following the first micturitionevent, and if after approximately 20 seconds no micturition event hadoccurred, then the bladder was emptied. However, if another micturitionevent occurred, the bladder was emptied after the bladder pressurereturned to baseline (FIG. 4A). Voided and residual volumes wererecorded and used to calculate bladder capacity and voiding efficiency.For saline-only experiments in which a second micturition event occurredbefore bladder emptying, the voided volume from the first and secondmicturition events were collected separately. For these trials, voidingefficiency was calculated as the first voided volume divided by thebladder capacity, and “reflex” voiding efficiency was calculated as thesum of the two voided volumes divided by the bladder capacity.

For PGE2 experiments, following control CMGs with saline the bladder wasinfused continuously with 100 μM PGE2 solution for 1 hour. The bladderwas subsequently emptied, and CMGs recorded while filling the bladderwith 100 μM PGE2. Following baseline PGE2 trials, trials with sensorypudendal stimulation were interleaved with PGE2 trials withoutstimulation.

For saline-only experiments, sensory pudendal stimulation trialsfollowed baseline control trials. Following trials in which stimulationincreased bladder capacity (approximately greater than 20%),non-stimulation trials were run until the bladder capacity returned to asteady-state (FIG. 5A). Stimulation amplitudes (0.3 T, 0.6 T, and 1 T,n=9, and 1.5 T, n=6 for 1 Hz, 10 Hz and n=7 for 20 Hz) were tested fromlowest to highest due to concerns of higher-amplitude stimuli producinglasting changes in bladder capacity. At each amplitude three stimulusrates were presented (1, 10, and 20 Hz) in randomized order. Stimuliwere biphasic, 100 μs per phase pulses delivered via a voltage tocurrent convertor (Model 2200, A-M Systems) controlled via an analogoutput channel on the PowerLab system. Electrical stimulation started atthe start of the pump and stopped at the first micturition event (n=9,PGE2 and n=7, saline-only) or went throughout the entire trial (n=2,saline-only).

For PGE2 experiments, stimulation testing started at 100 μA (n=1), 200μA (n=6), or 400 μA (n=2), all at 20 Hz. In cases where the stimulationdid not clearly increase bladder capacity (n=2), the stimulus amplitudewas increased by a factor of two until an increase in bladder capacitywas evident. In one experiment this occurred after one increase (2×),and in the other experiment two increases were necessary (4× initialamplitude).

Data Analysis

All signals were collected using a PowerLab/16SP acquisition unit (ADInstruments, Colorado Springs, Colo.) in conjunction with Labchart Profor visualization (Versions 7 & 8, AD Instruments). All analysis wasperformed using Matlab (Mathworks, Natick, Mass.). Bladder capacity wascalculated as the sum of the residual and voided volumes. Voidingefficiency was calculated as the voided volume divided by the bladdercapacity. For saline-only experiments in which two voids were allowed,the output from both voids was summed to create a secondary measure ofvoiding efficiency (reflex voiding efficiency).

For display, pressure traces were low-pass filtered using a first-orderzero-phase Butterworth filter with a 5 Hz cutoff. EUS EMG was high passfiltered at 70 Hz using a first-order zero-phase Butterworth filter.

Summary data are presented as either mean±standard error or as boxplotsusing Matlab's boxplot( ) function. All repeated trials within anexperiment were averaged together to create a single value perexperiment. Voiding efficiency and bladder capacity values werenormalized to non-stimulation saline control trials. For saline-onlyexperiments, the normalization was made relative to non-stimulationtrial data captured prior to any stimulation. Normalized data wereincluded in two-way ANOVA analysis (anovan( ), Matlab). Paired t-testswere used for comparison of different conditions, with the exception ofcomparison to control values where a t-test compared whether or not acondition's mean was different than 1(ttest(x,y) and ttest(x,1),Matlab). Tests with p<=0.05 were considered to be statisticallysignificant.

Example 1—Effect of Sensory Pudendal Nerve Stimulation duringSingle-Fill Cystometry on Bladder Capacity

In order to assess the possible use of pudendal nerve stimulation in thetreatment of OAB, bladder capacity following pudendal nerve stimulationwas assessed in female Wistar rats. Here bladder capacity was shown toincrease following pudendal nerve stimulation (FIG. 3).

The increase in bladder capacity was dependent upon stimulationamplitude and repetition frequency (two-way ANOVA, p<0.001 for bothamplitude and frequency, n=9 for 0.3 T to 1 T, n=7 for 1.5 T, 20 Hz, andn=6 for 1.5 T, 1 and 10 Hz). The largest increase was at 1.5 T at 10 Hz,in which the average bladder capacity was 253% of the non-stimulatedcontrol trials. Stimulation at 10 Hz led to the largest increases inbladder capacity, but the relative increases in bladder capacity werenot different between 10 and 20 Hz except at 0.6 T (p<0.001) (FIG. 3D).Increasing stimulation amplitude from 1 T to 1.5 T resulted in anincrease in bladder capacity for 1 Hz and 20 Hz stimulation (p=0.01 andp=0.03 respectively), but not for 10 Hz.

There was evidence of saturation at 10 Hz, as the increase in bladdercapacity was not statistically different between 1 T and 1.5 T. This wasdependent on the experiment, as can be seen by the differences inbladder capacity between 1 T and 1.5 T in FIG. 3A and FIG. 3B.

Example 2—Effect of Sensory Pudendal Nerve Stimulation duringSingle-Fill Cystometry on Voiding Efficiency

Further tests were performed in order to determine the effect of sensorypudendal nerve stimulation on voiding efficacy.

Increasing stimulation amplitude also led to a decrease in voidingefficiency. Changes in voiding efficiency were also dependent uponstimulation amplitude and repetition frequency (two-way ANOVA, p<0.001for both amplitude and frequency, n=7 for 0.3 T to 1 T, n=6 for 1.5 T,20 Hz, and n=5 for 1.5 T, 1 and 10 Hz) (FIG. 4B). On average 10 Hzstimulation led to the largest decreases in voiding efficiency. Athigher amplitudes, micturition events followed termination of thestimulus. However, at the amplitude which yielded the largest increasesin bladder capacity, “reflex” voiding efficiency was still less thancontrol values for 1 or 10 Hz (p=0.002 and p<0.001 respectively). Thereflex voiding efficiency was not statistically different from controlvalues for 1.5 T, 20 Hz.

At the highest stimulation amplitudes tested, it was not uncommon forsmall amounts of leaking to occur at high bladder pressures, rather thana coordinated contraction. Termination of the stimulus allowedsubsequent voids to occur, but voiding efficiency was still reducedrelative to non-stimulated trials.

Example 3—Effect of Sensory Pudendal Nerve Stimulation duringSingle-Fill Cystometry on Subsequent Non-Stimulated Trials

In order to determine whether the therapeutic effect of sensory pudendalnerve stimulation could provide an ongoing treatment for OAB once nervestimulation had been halted, the effect of sensory pudendal nervestimulation during single-fill cystometry on subsequent non-stimulatedtrials was investigated.

Stimulation of the sensory pudendal nerve often produced a carry-overeffect and bladder capacity was increased on subsequent non-stimulationtrials (FIG. 5A, B). Due to high variability in the magnitude of thiseffect, bladder capacity values from trials that followed 1 or 1.5 T and10 or 20 Hz were combined in summary data (FIG. 5C). Only groups oftrials in which stimulation increased bladder capacity by at least 20%relative to the preceding control trial and in which at least threenon-stimulated trials followed the stimulated trial were included (n=15,48% of trials). For the first trial following a stimulation trial,bladder capacity remained elevated by 55% (median) relative to baseline.For the third non-stimulation trial, with a void at an average of 39.5minutes following termination of the stimulus, the bladder capacityremained elevated by 29%.

The time to return back to a steady-state bladder capacity varied, buttended to occur within 1 hour. The utility of this observation isunclear, especially given that the maximum level of inhibition (maximumbladder capacity) occurred during stimulation, not after, but it maysuggest that intermittent stimulation will be sufficient to treat thesymptoms of OAB.

Example 4—Effect of Intravesical PGE2 on Bladder Capacity and VoidingEfficiency

Intravesical PGE2 reduces bladder capacity in rats (Ishizuka et al.1995) and causes “a strong urgency sensation” when given to healthywomen, also leading to reduced bladder capacities (Schüssler et al.1990). Consistent with these observations, intravesical PGE2 reducedbladder capacity.

Intravesical PGE2 decreased bladder capacity (p=0.02) and increasedvoiding efficiency (p<0.001, n=9) (FIG. 6). Sensory pudendal nervestimulation at 20 Hz increased bladder capacity relative to PGE2(p=0.004). Similar to the results with saline, an increase in bladdercapacity from stimulation also resulted in a decrease in voidingefficiency relative to the PGE2 condition. These preliminary resultsprovide evidence that sensory pudendal nerve stimulation may work undera variety of conditions.

All PGE2 experiments were conducted prior to saline-only experiments.The impact of intravesical PGE2 on bladder capacity appeared to changeover time. By comparison, intravesical saline-only appeared to be muchmore stable (consistent bladder capacity over time). Coupled with theunexpected presence of stimulation carryover effects which meant singletesting blocks could take 2-3 hours, saline-only was judged to be morewell-suited for initial exploration of the stimulus parameter space.

Sensory pudendal nerve stimulation increased bladder capacity in theanesthetized female Wistar rat during intravesical saline and PGE2conditions. 10 Hz stimulation at 1.5 T led to the largest increase inbladder capacity. Stimulation that produced increases in bladdercapacity also led to decreases in voiding efficiency.

Example 5—Effects of State-Dependent Pudendal Nerve Stimulation

In order to evaluate the effect of stimulating the pudendal nerve duringdifferent phases of the micturition cycle—also referred to as“state-dependent stimulation”—three experiments were conducted inalpha-chloralose anesthetized male cats.

Nerve cuff electrodes were placed on the sensory and motor branches ofthe pudendal nerve, just distal to the branching of the compoundpudendal nerve (pudendal sensory branch used was the dorsal nerve of thepenis (DNP)). Stimulation and control trials were interleaved withrandom within-block ordering.

In experiment 1, unstimulated controls (FIG. 7, trace A) were comparedto 3 stimulation regimes (FIG. 7, traces B-D). Stimulation (shown bydashed line below each trace) consisted of the stimulus during bladderfilling (3 T, 10 Hz), followed by either: B) the same stimulus duringvoiding (state independent); C) a termination of the stimulus at voidonset, D) switching the stimulation rate to 33 Hz at void onset. In asecond and third experiment, a fourth stimulation regime (E) was alsoused (FIGS. 8 and 9). In this regime, DNP stimulation was terminated atvoid onset and motor branch stimulation starting during voiding. Thestimulation pattern used during motor branch stimulation consisted ofstimulus trains at 40 Hz lasting for 100 ms in duration, repeated at 2Hz. Amplitudes of 1.8-2.3 T were selected as they maximized evokedexternal anal sphincter EMG. Approaches C-E are referred to asstate-dependent stimulation, as the stimulus parameters used depend onthe state of the subject—i.e. whether bladder filling or bladder voidingis occurring.

Average bladder capacity and voiding efficiency values are from 2-3cystometric trials.

Results

By using state-dependent stimulation it is shown it is possible to notonly increase bladder capacity beyond the no-stimulation condition, butthat voiding efficiency can be increased to higher levels than thoseobserved during state-independent stimulation.

In the first experiment, stimulation of the DNP throughout an entirecystometric trial (filling and voiding) increased bladder capacity (FIG.7B) relative to no-stimulation (FIG. 7A). By terminating the stimulus orincreasing the frequency of the stimulus signal at void onset voidingefficiency increased even further than when stimulation occurredthroughout the trial (state-dependent stimulation, FIG. 7C, D).

Similar results were obtained in the second experiment (FIG. 8). Inaddition to the previously tested approaches to state-dependentstimulation, the motor “bursting” pattern was tested during voiding(FIG. 8E) which led to increased voiding efficiency (50.8 ml, 83%, FIG.8E). In the third experiment, the baseline bladder capacity(no-stimulation condition) was elevated (compared to the other twoexperiments). Stimulation increased the bladder capacity, but onlyslightly (7% increase). In this experiment state-dependent stimulationwith motor bursting led to both an increase in bladder capacity andvoiding efficiency (FIG. 9E).

In three experiments state-dependent stimulation increased bladdercapacity relative to controls and voiding efficiency relative tostate-independent stimulation. In all experiments, stimulation of thesensory pudendal nerve increased bladder capacity. In two out of threeexperiments state-dependent stimulation using stimulus termination orswitching to higher frequency stimulation at onset of voiding led toincreased voiding efficiency. In two experiments state-dependentstimulation using motor branch burst stimulation during voidingincreased voiding efficiency.

II. Experimental Dataset 2

The following experimental methods were used throughout Examples 6-8discussed below.

Rat Surgical Preparation and Procedures

Female Wistar rats (n=25) weighing between 237 and 296 g wereanesthetized with urethane (1.2 g/kg SC, supplemented as necessary).Body temperature was monitored using an esophageal temperature probe andmaintained at 36-38° C. with a water blanket. Heart rate and arterialblood oxygen saturation levels were monitored using a pulse oximeter(Nonin Medical Inc., 2500 A VET).

In preparation for cystometrogram (CMG) measurements, the bladder wasexposed via a midline abdominal incision. The tip of a polyethylene(PE-90) catheter (Clay Adams, Parsippany, N.J.) was heated to create acollar and inserted into the bladder lumen through a small incision inthe apex of the bladder dome which was secured with a 6-0 silk suture.The abdominal wall was closed in layers with 3-0 silk suture. Thebladder catheter was connected via a 3-way stopcock to an infusion pump(Braintree Scientific Inc., BS-8000 or Harvard Apparatus PHD 4400) andto a pressure transducer (ArgoTrans, ArgonMedical Devices Inc., Plano,Tex.) connected to a bridge amplifier and filter (13-6615-50, GouldInstruments, Valley View, Ohio) for measuring intravesical pressure(IVP). Data were sampled at 1 kHz using a PowerLab system (ADInstruments, Colorado Springs, Colo.).

External urethral sphincter (EUS) EMG was measured using two platinumcontacts bonded to a silicone backing with wires welded to each contact(Microleads, Boston). This sheet-electrode was placed intra-abdominallybetween the urethra and the pubic symphysis. (Hokanson et al., 2017b).EUS EMG leads were connected through a preamplifier (HIPS, GrassProducts, Warwick, R.I.) to an amplifier (P511, Grass Products). Asubcutaneous needle served as ground. Signals were filtered (3 Hz-3 kHz)and sampled at 20 kHz.

After placing the bladder catheter and EUS EMG electrodes, the animalwas turned to a prone position for cuff placement. After resectinggluteal muscles at the midline, the ischium was spread apart from thesacrum to expose the ischiorectal fossa, and the sensory branch of thepudendal nerve was isolated from connective tissue. A custom nerve cuffwas used in one experiment. In other experiments either a 300 μm innerdiameter with 3 mm length (n=6) or 200 μm inner diameter (2 mm length,n=18) were used (CorTec, Freiburg, Germany). In experiments with motorbranch stimulation, the ischiorectal fossa on the opposite side of theanimal was exposed and the pudendal motor nerve and blood vessels wereisolated. Due to the small size of the motor nerve branch, a 400 μminner diameter cuff was placed around the pudendal motor branch as wellas the pudendal blood vessels (CorTec, n=9). Following nerve cuffplacement, the incision was sutured closed in layers with 3-0 silksuture. The animal was then turned back into a supine position forcystometric testing.

The gross neural anatomy of the lower urinary tract in the rat is shownin FIG. 10A. The left inset shows this anatomy relative to the pelvicbones and spinal column. The area highlighted in the inset correspondsto our access point, in the ischiorectal fossa. A picture of thedissection of the sensory nerve branch is shown in FIG. 10B, with a 300μm inner diameter, 3 mm length positioned close to, but not yet around,the nerve. The type of cuff used in these experiments is shown in FIGS.10D and 10E. The cuff is inserted around the nerve by pulling on twotabs which open the cuff. Releasing the tabs closes the cuff.

Rat Electrical Stimulation

Electrical stimulation was delivered using a stimulus isolator (n=18) ora stimulus generator (n=7, model STG4002-16 mA, Multi-Channel Systems,Reutlingen, Germany). Stimulation pulses consisted of a charge-balancedbiphasic waveform with 100 μs pulse widths. Strength of stimulation wasassessed by monitoring evoked EUS EMG. Amplitudes for sensory pudendalnerve stimulation were normalized to the minimum stimulation amplitudenecessary to reflexively evoke EUS EMG activity. This stimulationamplitude is referred to as 1 T (1 times threshold amplitude). Motorbranch stimulation occurred at the minimum stimulus amplitude requiredto evoke a maximal (direct) EUS EMG response.

Electrical stimulation to promote bladder filling started at the onsetof bladder filling. For one set of animals (n=11), the inhibitorypattern consisted of stimulation at either 1, 10 or 20 Hz and atstimulation amplitudes of 0.3, 0.6, 1, and 1.5 T. In these experimentsstimulation persisted throughout the first bladder contraction which ledto leakage or a void. An additional one minute worth of data wascollected to see termination of the stimulus led to a reflexive voidwhich increased the voiding efficiency back to or above control values.In a second set of experiments (n=9) exploring state-dependentstimulation, stimulation occurred at 10 Hz and 1 T. Depending on thetype of trial, stimulation either continued throughout bladder emptyingor terminated just prior to bladder emptying. All stimulation to promotebladder filling occurred on the sensory pudendal nerve.

Electrical stimulation to promote bladder emptying started just prior tobladder voiding or urine leakage and continued throughout the bladdercontraction. In the first set of state-dependent stimulation experiments(n=9), four approaches to promoting bladder emptying were employed. Forthe first approach no stimulation occurred, but rather the inhibitorystimulus was terminated such that no stimulation was occurring duringbladder emptying. The other three approaches, two of which are shown inFIG. 10F, consisted of three pulses at 40 Hz repeated at either 2 Hz(every 0.5 seconds), 4.76 Hz (every 0.21 seconds), or 8 Hz (every 0.125seconds). These stimulation patterns were all delivered on the motorbranch and we refer to these patterns as motor “bursting” patterns.

Cat Surgical Preparation and Procedures

Acute experiments were conducted in adult neurologically intact male(n=6, 3.4-3.8 kg) and female (n=5, 2.8-3.2) cats. Anesthesia was inducedwith isoflurane (3%) and maintained with α-chloralose (65 mg/kg initialdose followed by continuous infusion of 5 mg/kg/h iv and supplemented asnecessary based on jaw tone and blood pressure) following completion ofthe surgery. Gentamicin 5 mg IM and ketofen 1.2 g/kg SQ were given priorto surgical incision. A tracheotomy was performed to place a siliconeendotube (Cat. no J0612B, Jorgensen Laboratories, Loveland, Colo.)connected to an artificial respirator (ADS 1000, Engler EngineeringCorporation, Hialeah, Fla.), and artificial respiration was controlledto maintain end-tidal CO₂ between 3-4% (Capnogaurd, Novametrix MedicalSystems Inc., Wallingford, Conn.). The right carotid artery wascannulated with a 3.5 Fr polypropylene catheter (Cat. no 8890703211,Medtronic, Minneapolis, Minn.) to monitor arterial blood pressure(Tektronix 413A Neonatal Monitor) and was kept patent by infusing salineat a constant rate of 6 ml/hr. Body temperature was measured using anesophageal temperature probe and maintained at 38° C. with a forced-airwarming blanket (Bair Hugger model 505, 3M). Additional fluids (0.9%physiological saline with 5 dextrose and 8.4 g/L NaHCO₃) wereadministered continuously (15 ml/kg/hr, i.v.) via the left cephaliccatheter. Following a midline abdominal incision, the bladder wascannulated through the dome with a modified 14 g BD Angiocath catheterconnected to PE 90 tubing introduced with a hypodermic needle, securedwith a purse string suture (4-0 silk, Cat. No M-S418R19, AD Surgical,Sunnyvale, Calif.) and connected to a solid-state pressure transducer(Deltran, Utah Medical, Utah) to measure bladder pressure. A forcetransducer (model: MLT500D, AD Instruments, Colorado Springs, Colo.) wasused to collect voided volume (VV). The external anal sphincter (EAS)EMG activity was measured by PFA-coated platinum-iridium wires (0.0055inch-diameter, A-M Systems, Sequim, Wash.) inserted percutaneously intothe EAS bilaterally. EAS EMG leads were connected through a preamplifier(HIPS, Grass Products, Warwick, R.I.) to an amplifier (P511, GrassProducts). Bladder pressure (BP), VV, and EAS EMG signals wereamplified, filtered, and sampled at either 1,000 Hz (BP and VV) or20,000 Hz (EAS EMG).

The bladder was continuously filled with physiological saline at roomtemperature (0.2-4 ml/min, median=1.3 ml/min) using an infusion pump(model: PHD 4400, Harvard Apparatus), with an open urethra forapproximately one hour to allow post-surgical recovery. The bladder wassubsequently emptied and cystometrograms (CMGs) recorded. For each CMG,the bladder was filled micturition or urine leakage occurred until, atwhich time the infusion pump was turned off. Approximately one minuteafter the bladder pressure returned to baseline, the bladder was emptiedvia the catheter using a syringe. Within a block of trials, the fillingrate remained constant. Voided (VV) and residual (RV) volumes wererecorded and used to calculate bladder capacity (BC) and voidingefficiency (VE).

Cat Electrical Stimulation

Electrical stimulation was delivered using a stimulus generator (modelSTG4002-16 mA, n=4 and model STG4004-16 mA, n=7, Multi-Channel Systems).Stimulation pulses consisted of a charge-balanced biphasic waveform with100 μs per phase. Strength of stimulation was assessed by monitoringevoked EAS EMG. For sensory pudendal stimulation (females) and dorsalgenital nerve stimulation (males), stimulus threshold was defined as thestimulus amplitude necessary to evoke reflex EAS EMG activity. For motorbranch (i.e. rectal-perineal nerve) nerve stimulation, the minimumamplitude that evoked a maximal EAS EMG response was used.

Electrical stimulation to promote bladder filling started at fillingonset. The stimulus consisted of sensory pudendal nerve (female) ordorsal genital nerve (males) stimulation at 3 T and 10 Hz. In sometrials this stimulation persisted throughout a voiding contraction.

In other trials state-dependent stimulation was employed in an attemptto increase voiding efficiency. One approach for state-dependentstimulation was to terminate the stimulus at void onset. A secondapproach was to change the stimulation rate on the sensory pudendal ordorsal genital nerves from 10 Hz to 33 Hz at void onset. The thirdapproach was to employ a bursting pattern at void onset. All burstingpatterns consisted of 3 pulses at 40 Hz at a 2 Hz train rate. Unlike therat experiments the train rate was not changed. For motor stimulation,this bursting occurred at the minimum stimulus amplitude which generatedthe maximal EAS EMG response. For sensory pudendal or dorsal genitalnerve stimulation this occurred at 3 T, the same amplitude used for 10Hz stimulation during bladder filling.

Data Analysis

For each trial (cystometrogram) bladder capacity was calculated as thesum of the voided volume and the residual volume extracted from thebladder. Voiding efficiency was calculated as the ratio of the voidedvolume to the bladder capacity.

Stimulus patterns were randomized within block and values werenormalized to preceding control levels for plotting and summarystatistics (e.g. median values). For state-dependent stimulation datavoiding efficiency was compared between no-stimulation controls,continuous stimulation, and the state-dependent stimulation conditions.This was done by use of Friedman ANOVA test, followed by the Benjamini,Krieger and Yekutieli two-stage step-up method for post-hoc comparisonof conditions to continuous stimulation. This ANOVA test requires nomissing data for any stimulus condition, so the number of complete datasets was reduced to five for both state-dependent stimulation in malecats and female rats. The ANOVA and post-hoc testing were computed usingGraphPad (Version 7.04, La Jolla, Calif.).

Example 6—Impact of Continuous Sensory Pudendal Stimulation on BladderCapacity as a Function of Stimulation Amplitude and Pulse RepetitionRate

In female rats (n=9) the impact of continuous sensory pudendalstimulation on bladder capacity was evaluated as function of stimulationamplitude and pulse repetition rate. Example cystometrograms from oneanimal are shown in FIG. 11A. Stimulation continued until urine was losteither through leakage or voiding contraction. Increasing stimulationamplitude increased bladder capacity and the increase in bladdercapacity was dependent on stimulation rate (FIG. 11 B). The largestincreases in bladder capacity came from stimulation at 1.5 T, withaverage normalized bladder capacities of 1.65 (1 Hz), 2.53 (10 Hz), and2.25 (20 Hz) of control values. In addition to increasing bladdercapacity, increasing stimulation amplitude also decreased voidingefficiency. This can also be seen in the pressure traces (FIG. 11A)where increasing stimulation amplitude led to slow rises in bladderpressure towards the end of filling, rather than coordinatedcontractions.

Example 7—Continuous Stimulation of the Dorsal Genital Nerve (DGN) andState-Dependent Stimulation in Male Cats

FIG. 12 shows the results of experiments using continuous stimulation ofthe dorsal genital nerve (DGN) and state-dependent stimulation as testedin male cats (n=6). In this example continuous stimulation decreasedvoiding efficiency by 24%. Across all experiments continuous stimulationdecreased voiding efficiency by 33% (median) relative to controls (FIG.12C), confirming the results from experiments in rats, although thistrend was not statistically significant (p=0.072, n=5).

Bladder capacity results were combined across all stimulation trials asstimulation during bladder filling was identical in all trials up untilvoid onset and stopping of the infusion pump. All experiments showedincreased bladder capacity from DGN stimulation with a median increaseof 26% relative to the no-stimulation condition (FIG. 12B).

Both termination of continence-promoting inhibitory stimulation at theonset of voiding, and 33 Hz stimulation during voiding on averageincreased voiding efficiency relative to control voiding efficiency withno inhibitory stimulation (14% and 19% medians respectively), althoughneither was statistically significant (p>0.05, n=5). The largest andmost consistent increase in VE came from switching to burst stimulationof the motor pudendal nerve branch, which produced a 379% medianincrease in VE (p=0.028). Relative to continuous stimulation,state-dependent stimulation led to statistically-significant increasesin voiding efficiency (p=0.003 for ANOVA test, p=0.028 for fill only,p=0.048 for 33 Hz, p<0.001 for motor bursting, n=5) (data shown in FIG.12C includes n=5 for motor bursting, n=6 for others).

Similar data was collected in female cats (n=5). Example traces from anexperiment are shown in FIG. 14A. Continuous stimulation increasedbladder capacity but decreased voiding efficiency by 52%. Side-by-sidemale and female cat summary data is shown in in FIGS. 14B and 14C.State-dependent stimulation increased bladder capacity in males by 26%(median, n=6 males) and in females 15% (median, n=5) relative to controltrials. In female cats continuous stimulation decreased voidingefficiency (median 21% decrease). Unlike in male cats, fill-only and 33Hz stimulation conditions yielded voiding efficiencies that are onaverage less than controls (42% and 50% decreases respectively relativeto controls) in female cats. Similar to male cats, state-dependentstimulation in female cats with motor bursting increased bladdercapacity and voiding efficiency (median 100% increase) relative to thecontrol condition. Interestingly, in all experiments, motor burstingincreased voiding efficiency relative to control trials.

Example 8—State-Dependent Stimulation using Motor Bursting to IncreaseVoiding Efficiency in the Rat

FIG. 13 shows results of state-dependent stimulation using motorbursting to increase voiding efficiency in the rat. Termination of thestimulus, and in some trials, a transition to motor bursting occurredjust prior to the volume at which a bladder contraction was anticipated.As expected and consistent with previous experimental results (FIG. 11),sensory pudendal nerve stimulation increased bladder capacity to 158%(median) of no-stimulation control trials (FIG. 13B, n=9). Statedependent stimulation also increased voiding relative to continuousstimulation (data shown in FIG. 13C includes n=5 for 8 Hz bursting, n=8for 4.76 Hz bursting, n=9 for others) (p=0.023 for ANOVA, p=0.016 forfill-only, p=0.001 for 2 Hz, p=0.003 for 4.76 Hz, p=0.004 for 8 Hz,n=5). Median normalized voiding efficiency values were 19% (continuousstimulation), 48% (fill-only), 98% (2 Hz motor), 117% (4.76 Hz motor),and 75% (8 Hz motor).

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Clauses

The following clauses describe further embodiments of the invention:

Clause 1. An apparatus for stimulating neural activity in a pudendalnerve of a subject, the apparatus comprising:

at least one primary electrode configured to apply a first electricalsignal to said nerve; and a controller coupled to said primaryelectrode(s) and controlling the first electrical signal to be appliedthereby,

wherein said controller is configured to cause said at least one primaryelectrode to apply said first electrical signal that stimulates neuralactivity in the pudendal nerve to produce an increase in bladdercapacity, wherein the first electrical signal comprises an AC waveformhaving a frequency in the range of from 1-50 Hz and wherein the firstelectrical signal has an amplitude in the range from 0.1 to 10 T.

Clause 2. An apparatus according to clause 1 wherein the firstelectrical signal has an amplitude in the range from 0.3 T to <2.0 T.

Clause 3. An apparatus according to clause 1 or clause 2 wherein thefirst electrical signal has an amplitude in the range from 1 T to 1.5 T.

Clause 4. An apparatus according to any one of clauses 1-3 wherein thefirst electrical signal comprises an AC waveform having a frequency inthe range of from 1-20 Hz.

Clause 5. An apparatus according to any one of clauses 1-4 wherein thefirst electrical signal comprises an AC waveform having a frequency of10 Hz or 20 Hz.

Clause 6. An apparatus according to clause 1 wherein the firstelectrical signal has an amplitude in the range from 2 T to 10 T.

Clause 7. An apparatus according to clause 6 wherein the firstelectrical signal has an amplitude in the range from 2 T to 4 T,optionally 3 T.

Clause 8. An apparatus according to clause 6 or clause 7 wherein thefirst electrical signal comprises an AC waveform having a frequency inthe range of from 1-20 Hz.

Clause 9. An apparatus according to clause 8 wherein the firstelectrical signal comprises an AC waveform having a frequency of 10 Hz.

Clause 10. An apparatus according to any one of clauses 1 to 9, whereinsaid at least one primary electrode is configured to apply said firstelectrical signal to sensory fibres of said pudendal nerve, and saidcontroller is configured to cause said at least one primary electrode toapply said first electrical signal that stimulates neural activity insensory fibres of the pudendal nerve to produce an increase in bladdercapacity.

Clause 11. An apparatus according to any one of clauses 1-10, whereinsaid controller is configured to cause a second electrical signal to beapplied, wherein said second electrical signal stimulates neuralactivity in the pudendal nerve to produce an increase in voidingefficiency and wherein the second electrical signal comprises an ACwaveform having a frequency higher than the frequency of the firstelectrical signal.

Clause 12. An apparatus according to clause 11, wherein said secondelectrical signal is applied by said at least one primary electrode(s).

Clause 13. An apparatus according to clause 11, wherein said secondelectrical signal is applied by at least one secondary electrode(s)coupled to said controller, said controller controlling the signal to beapplied thereby.

Clause 14. An apparatus according to any one of clauses 1-10, furthercomprising at least one secondary electrode configured to apply a secondelectrical signal to said nerve and coupled to said controller, saidcontroller controlling the signal to be applied thereby, wherein saidcontroller is configured to cause said secondary electrode to apply saidsecond electrical signal that stimulates neural activity in the pudendalnerve to produce an increase in voiding efficiency, wherein the secondelectrical signal comprises an AC waveform and wherein said controlleris configured to cause said second electrical signal to be applied in aburst pattern.

Clause 15. An apparatus according to clause 14, wherein said burstpattern consists of a signal burst having a duration from 50 ms to 1000ms repeated at an interval of from 0.2 s to 2 s.

Clause 16. An apparatus according to clause 14 or 15, wherein said burstpattern consists of a signal burst having a duration of 100 ms repeatedat an interval of 0.5 s.

Clause 17. An apparatus according to any one of clauses 11-16, whereinthe second electrical signal comprises an AC waveform having a frequencyin the range of from 20-50 Hz.

Clause 18. An apparatus according to clause 17 wherein the secondelectrical signal comprises an AC waveform having a frequency of 30-40Hz, optionally 33 Hz or 40 Hz.

Clause 19. An apparatus according to any one of clauses 11-18 whereinthe second electrical signal comprises an AC waveform has an amplitudein the range of 0.5-4 T, optionally 1-3 T.

Clause 20. An apparatus according to clause 14-19, wherein said at leastone secondary electrode is configured to apply said second electricalsignal to motor fibres of said pudendal nerve, and said controller isconfigured to cause said at least one secondary electrode(s) to applysaid second electrical signal that stimulates neural activity in motorfibres of the pudendal nerve to produce an increase in voidingefficiency.

Clause 21. An apparatus according to any one of clauses 1 to 20, whereinsaid primary electrode(s) is a bipolar cuff electrode.

Clause 22. An apparatus according to any one of clauses 13-21, whereinsaid secondary electrode(s) is a bipolar cuff electrode.

Clause 23. An apparatus according to any one of clauses 1-22, whereinthe apparatus further comprises a detector to detect one or morephysiological parameters in the subject, wherein the controller iscoupled to said detector, and causes the electrical signal to be appliedwhen a physiological parameter is detected to be meeting or exceeding apredefined threshold value.

Clause 24. An apparatus according to clause 23, wherein one or more ofthe detected physiological parameters is selected from nerve activity inthe pudendal nerve, nerve activity in the hypogastric nerve, nerveactivity in the pelvic nerve, muscle activity in the bladder detrusormuscle, muscle activity in the internal urethral sphincter, muscleactivity in the external urethral sphincter, muscle activity in theexternal anal sphincter, and bladder pressure.

Clause 25. An apparatus according to any one of clauses 23 or 24,wherein the controller causes the first electronic signal to be stoppedwhen the detector detects onset of a bladder voiding phase.

Clause 26. An apparatus according to any one of clauses 1-25, whereinthe controller is configured to cause the first electrical signal to beapplied no more frequently than alternate micturition cycles, optionallyno more frequently than every third micturition cycle.

Clause 27. An apparatus according to any one of clauses 1-26, furthercomprising an input element, wherein the input element allows thesubject to enter data regarding their behaviour and/or desires so as todetermine when the controller causes the electrical signal to beapplied.

Clause 28. An apparatus according to clause 27, wherein the inputelement is in wireless communication with the controller.

Clause 29. An apparatus according to any one of clauses 1-28 wherein theapparatus is suitable for at least partial implantation into thesubject, optionally full implantation into the subject.

Clause 30. A method of treating bladder dysfunction in a subjectcomprising:

i. implanting in the subject an apparatus according to any one ofclauses 1-29;

ii. positioning at least one primary electrode of the apparatus insignalling contact with a pudendal nerve of the subject and, when theapparatus comprises at least one secondary electrode, positioning saidat least one secondary electrode of the apparatus in signalling contactwith a pudendal nerve of the subject;

iii. activating the apparatus to apply an electrical signal to thepudendal nerve of the subject as caused by the controller.

Clause 31. A method of treating bladder dysfunction in a subjectcomprising applying a first electrical signal to a pudendal nerve of thesubject, wherein the first electrical signal comprises an AC waveformhaving a frequency in the range of from 1-50 Hz and wherein the firstelectrical signal has an amplitude in the range of from 0.1 to 10 T.

Clause 32. A method according to clause 31, wherein said firstelectrical signal is applied to the pudendal nerve during a bladderfilling phase, wherein application of said first electrical signalincreases bladder capacity.

Clause 33. A method according to any one of clauses 31-32 whereinapplication of said first electrical signal is stopped at the onset of abladder voiding phase.

Clause 34. A method according to any one of clauses 31-33, wherein thefirst electrical signal has an amplitude in the range from 0.3 T to <2.0T.

Clause 35. A method according to clause 34 wherein the first electricalsignal has an amplitude in the range from 1 T to 1.5 T.

Clause 36. A method according to any one of clauses 31-35, wherein thefirst electrical signal comprises an AC waveform having a frequency inthe range of from 1-20 Hz.

Clause 37. A method according to clause 36 wherein the first electricalsignal comprises an AC waveform having a frequency of 10 Hz or 20 Hz.

Clause 38. A method according to any one of clauses 31-33 wherein thefirst electrical signal has an amplitude in the range of from 2 T to 10T.

Clause 39. A method according to clause 38 wherein the first electricalsignal has an amplitude in the range from 2 T to 4 T, optionally 3 T.

Clause 40. A method according to clause 38 or clause 39 wherein thefirst electrical signal comprises an AC waveform having a frequency inthe range of from 1-20 Hz.

Clause 41. A method according to clause 40 wherein the first electricalsignal comprises an AC waveform having a frequency of 10 Hz.

Clause 42. A method according to any one of clauses 31-41, furthercomprising applying a second electrical signal to a pudendal nerve ofthe subject, wherein the second electrical signal comprises an ACwaveform having a frequency higher than the frequency of the firstelectrical signal.

Clause 43. A method according to any one of clauses 31-41, furthercomprising applying a second electrical signal to a pudendal nerve ofthe subject, wherein the second electrical signal comprises an ACwaveform and is applied in a burst pattern.

Clause 44. A method according to clause 43, wherein said burst patternconsists of a signal burst having a duration from 50 ms to 1000 msrepeated at an interval of from 0.125 s to 2 s.

Clause 45. A method according to clause 44, wherein said burst patternconsists of a signal burst having a duration of 100 ms repeated at aninterval of 0.5 s.

Clause 46. A method according to any one of clauses 42-45, wherein saidsecond electrical signal is applied to the pudendal nerve during abladder voiding phase, wherein application of said second electricalsignal promotes bladder voiding.

Clause 47. A method according to any one of clauses 42-46, wherein thesecond electrical signal comprises an AC waveform having a frequency inthe range of from 20-50 Hz.

Clause 48. A method according to clause 47 wherein the second electricalsignal comprises an AC waveform having a frequency of 30-40 Hz,optionally 33 Hz or 40 Hz.

Clause 49. A method according to any one of clauses 42-48 wherein thesecond electrical signal comprises an AC waveform has an amplitude inthe range from 0.5-4 T, optionally from 1-3 T.

Clause 50. A method according to any one of clauses 43-49, wherein thesecond electrical signal is applied to motor fibres of the pudendalnerve.

Clause 51. A method according to any one of clauses 30-50, wherein thefirst electronic signal is applied bilaterally and, in a methodaccording to clause 31, the apparatus comprises two primary electrodesand step (ii) comprises positioning the primary electrodes bilaterally.

Clause 52. A method according to any one of clauses 30, and 42-51,wherein the second electronic signal is applied bilaterally and, in amethod according to clause 30, the apparatus comprises two secondaryelectrodes and step (ii) comprises positioning the secondary electrodesbilaterally.

Clause 53. A method according to any one of clauses 31-52, wherein thefirst electronic signal is applied no more frequently than alternatemicturition cycles, optionally no more frequently than every thirdmicturition cycle.

Clause 54. A pharmaceutical composition comprising a compound fortreating bladder dysfunction, for use in a method of treating bladderdysfunction in a subject, wherein the method is a method according toany one of clauses 30-53, the method further comprising the step ofadministering an effective amount of the pharmaceutical composition tothe subject.

Clause 55. A pharmaceutical composition comprising a compound fortreating bladder dysfunction, for use in treating bladder dysfunction ina subject, the subject having an apparatus according to any one ofclauses 1-29 implanted.

Clause 56. A pharmaceutical composition for use according to clause 54or 55, wherein the compound for treating bladder dysfunction is anantimuscarinic compound or a β-adrenergic receptor agonist, optionally aβ3-adrenergic receptor agonist.

Clause 57. A pharmaceutical composition for use according to any one ofclauses 54-56, wherein the compound for treating bladder dysfunction isan antimuscarinic compound selected from darifenacin, hyoscyamine,oxybutynin, tolterodine, solifenacin, trospium, or fesoterodine.

Clause 58. A pharmaceutical composition for use according to any one ofclauses 54-56, wherein the compound for treating bladder dysfunction isa β3-adrenergic receptor agonist, optionally mirabegron.

Clause 59. A neuromodulation system comprising a plurality ofapparatuses according to any one of clauses 1-29.

Clause 60. A neuromodulation system according to clause 59, wherein eachapparatus is arranged to communicate with at least one other apparatusin the system, optionally all apparatuses in the system.

Clause 61. A neuromodulation system according to clause 59 or 60,further comprising a processor arranged to communicate with theapparatuses of the system.

1. An apparatus for stimulating neural activity in a pudendal nerve of asubject, the apparatus comprising: at least one primary electrodeconfigured to apply a first electrical signal to said nerve; and acontroller coupled to said primary electrode(s) and controlling thefirst electrical signal to be applied thereby, wherein said controlleris configured to cause said at least one primary electrode to apply saidfirst electrical signal that stimulates neural activity in the pudendalnerve to produce an increase in bladder capacity, wherein the firstelectrical signal comprises an AC waveform having a frequency in therange of from 0.1-50 Hz and wherein the first electrical signal has anamplitude in the range from 0.05 T to 10 T. 2-8. (canceled)
 9. Theapparatus according to claim 1 in which the first electrical signal isapplied no more frequently than alternate micturition cycles. 10-19.(canceled)
 20. The apparatus according to claim 1, in which at least thefirst electrical signal is applied to sensory fibres of the pudendalnerve.
 21. The apparatus according to claim 20 in which the signal isapplied to a sensory branch of the pudendal nerve.
 22. The apparatusaccording to claim 21 in which the at least one primary electrode isconfigured to apply the first electrical signal to the sensory branch ofthe pudendal nerve, and the controller is configured to cause said atleast one primary electrode to apply the first electrical signal thatstimulates neural activity in the sensory branch of the pudendal nerveto produce an increase in bladder capacity.
 23. The apparatus accordingto claim 1 in which application of the first electrical is stopped atonset of a bladder voiding phase.
 24. The apparatus according to claim23 in which the controller causes the first electrical signal to bestopped when onset of a voiding phase is detected.
 25. The apparatusaccording to claim 1 wherein said controller is configured to cause asecond signal to be applied wherein said second signal stimulates thepudendal nerve, wherein said second electrical signal stimulated neuralactivity in the pudendal nerve to procedure an increase in voidingefficiency and wherein the second electrical signal comprises an ACwaveform having a frequency in the range of from 20 to 50 Hz.
 26. Theapparatus according to claim 25 wherein the second electrical signal isapplied by said at least one primary electrode(s) or wherein theapparatus comprises at least one secondary electrode coupled to thecontroller and the second electrical signal is applied by the at leastone secondary electrode(s), the controller controlling the signal to beapplied thereby. 27-32. (canceled)
 33. The apparatus according to claim25 in which the second electrical signal is applied in a burst patterncomprising a signal burst comprising an AC waveform, wherein the signalburst is repeated at a frequency in the range of from 0.5-20 Hz. 34.(canceled)
 35. The apparatus according to claim 33 in which the burstpattern comprises a signal burst having a duration from 50 ms to 1000 msrepeated at an interval of from 0.125 s to 2 s. 36-38. (canceled) 39.The apparatus according to claim 25 in which the second electricalsignal is to be applied to motor fibres of the pudendal nerve.
 40. Theapparatus according to claim 39 in which the at least one secondaryelectrode(s) is (are) configured to apply the second electrical signalto motor fibres of said pudendal nerve, and the controller is configuredto cause said at least one secondary electrode to apply the secondelectrical signal that stimulates neural activity in motor fibres of thepudendal nerve to produce an increase in voiding efficiency. 41-49.(canceled)
 50. The apparatus according to claim 25 in which a secondelectrical signal is (to be) applied, the second electrical signalcomprises an AC waveform having an amplitude in the range of 0.5-4 T.51-54. (canceled)
 55. A method of treating bladder dysfunction in asubject comprising applying a first electrical signal to a pudendalnerve of the subject, wherein the first electrical signal comprises anAC waveform having a frequency in the range of from 1-50 Hz and whereinthe first electrical signal has an amplitude in the range of from 0.05 Tto 10 T. 56-62. (canceled)
 63. The method according to claim 55 in whichthe first electrical signal is applied no more frequently than alternatemicturition cycles. 64-73. (canceled)
 74. The method according to claim55 in which the first electrical signal is applied to sensory fibres ofthe pudendal nerve.
 75. The method according to claim 74 in which thesignal is applied to a sensory branch of the pudendal nerve.
 76. Themethod according to claim 55 in which application of the firstelectrical signal is stopped at onset of a bladder voiding phase. 77.(canceled)
 78. The method according to claim 55 further comprising theapplication of a second electrical signal to stimulate the pudendalnerve.
 79. (canceled)
 80. The method according to claim 78 in which thesecond electrical signal applied comprises an AC waveform having afrequency in the range from 20-50 Hz. 81-83. (canceled)
 84. The methodaccording to claim 78 in which the second electrical signal comprises anAC waveform and is applied in a burst pattern.
 85. The method accordingto claim 84 in which the burst pattern comprises a signal burst having aduration from 50 ms to 1000 ms.
 86. (canceled)
 87. The method accordingto claim 78 in which the second electrical signal is to be applied tomotor fibres of the pudendal nerve.
 88. The method according to claim 84in which the second electrical signal applied in a burst patterncomprises an AC waveform having a frequency in the range of from 20-50Hz. 89-90. (canceled)
 91. The method according to claim 84 in which thesecond electrical signal is applied in a burst pattern comprising asignal burst comprising the AC waveform, wherein the signal burst isrepeated at a frequency in the range of from 2 to 20 Hz. 92-96.(canceled)
 97. The method according to claim 84 wherein the burst patteris repeated at an interval of from 0.125 s to 2 s.
 98. (canceled) 99.The method according to claim 84 in which a second electrical signal is(to be) applied, wherein the second electrical signal comprises an ACwaveform having an amplitude in the range of 0.5-4 T. 100-103.(canceled)